CN109971848A - A grading model for detecting benign and malignant degree of esophageal tumor and/or gastric tumor and its application - Google Patents

Abstract

The present invention relates to a grading model for detecting the benign and malignant degree of esophageal tumor and/or gastric tumor and its application. The changes of imprinted genes in tumors are graded. The detection model and device of the present invention express the expression of imprinting deletion on tissue and cell samples of patients with esophageal tumors and gastric tumors in an intuitive way. The method can objectively, intuitively, early and accurately detect the changes of imprinted (trace) genes, and can provide a quantitative model, making a great contribution to the diagnosis of esophageal tumors and gastric tumors.

Description

A grading model for detecting the benign and malignant degree of esophageal and/or gastric tumors and its application

technical field

The invention relates to the field of biotechnology, to the field of gene diagnosis, to a grading model and application thereof, to a grading model for detecting the benign and malignant degree of esophageal tumor and/or gastric tumor and its application, in particular to a group of imprinted genes A grading model in detecting the benign and malignant degree of esophageal tumors and/or gastric tumors, and devices and applications of its components.

Background technique

Gastric cancer and esophageal cancer are two common digestive tract tumors. In 2012, there were 952,000 new gastric cancer cases and 723,000 deaths worldwide, 456,000 new esophageal cancer cases and 400,000 deaths. my country is a high-incidence area of gastric cancer and esophageal cancer. The new cases and deaths of gastric and esophageal cancer account for about 50% of the world each year. The 5-year survival rate of early gastric cancer and esophageal cancer exceeds 90%, while the 5-year survival rate of advanced gastric and esophageal cancer is less than 30%. Therefore, early diagnosis of gastric and esophageal cancer is of great significance to save the lives of patients. However, gastric cancer and esophageal cancer usually have no obvious symptoms in the early stage, and the detection rate in my country is less than 10%.

It has been recognized that intraepithelial neoplasia is a precancerous lesion of the stomach. According to the degree of hyperplasia, it can be divided into low-grade intraepithelial neoplasia and high-grade intraepithelial neoplasia. Low-grade intraepithelial neoplasia has an approximately 20% chance of developing gastric cancer, while high-grade intraepithelial neoplasia has a greater than 70% chance of developing gastric cancer. At present, the pathological distinction between low-grade and high-grade intraepithelial neoplasia is mainly subjective judgment by pathologists based on morphology. While several studies have attempted to remove the subjectivity of classification using mucous histochemistry, immunocytochemistry to detect differentiation or cell proliferation markers, and oncogene product and morphometric analysis, none has so far been more robust than traditional morphological analysis. obvious advantage. Similarly, the current diagnosis of esophageal cancer also mainly relies on imaging and histomorphology, and the accuracy of diagnosis largely depends on the experience and subjective judgment of doctors, which is prone to misdiagnosis.

CN 106609037 A discloses a detection kit for diagnosing gastric cancer patients based on multiple genes. The kit includes primers for diagnosing gastric cancer, which can be used for diagnosing gastric cancer, but the detection is not accurate enough to distinguish which specific gastric cancer is in stage.

The diagnosis of benign and malignant cells by traditional pathology is based on the size, morphology, infiltration and the relationship between the surrounding cells and tissues. It has great limitations in the discovery of early changes in cells (cancer), so the method for diagnosing cancer at the cellular and molecular level once became a research hotspot. With the continuous in-depth research in the field of molecular biology, more and more molecular detection technologies are applied to cancer diagnosis.

Cancer arises from uncontrolled cell growth/division caused by accumulation of epigenetic changes and genetic variations over time. The traditional pathological diagnosis is based on the variation in the size, shape and structure of cells and tissues, so as to make the judgment of benign and malignant of esophageal and gastric tumors. With the development and deepening of molecular biology, more and more molecular detection technologies have been applied to the detection of esophageal cancer and gastric cancer. From the analysis of the development process of cancer, changes at the molecular level (epigenetics and genetics) are far earlier than changes in cell morphology and tissue structure. Therefore, molecular biological detection is more sensitive for early detection of cancer.

Genomic imprinting is a form of gene regulation in epigenetics. It is characterized by methylation of alleles from a specific parent, so that only one allele of a gene is expressed, while the other is in a state of gene silencing. Genes of this type are called imprinted (marked) genes. Loss of imprinting is an epigenetic change in which demethylation of imprinted genes leads to the activation of silent alleles and the initiation of gene expression. Numerous studies have shown that this phenomenon (absence of imprinting) is prevalent in various types of cancer and occurs earlier than cellular and tissue morphological changes. At the same time, in healthy cells, the proportion of missing imprints was extremely low, in stark contrast to cancer cells. Therefore, the methylation status of imprinted genes can be used as a pathological marker to analyze the abnormal state of cells through specific molecular detection techniques.

Based on the above reasons, the current diagnosis of esophageal cancer and gastric cancer requires a new detection system and detection model, based on patient biopsy samples, to analyze the changes of molecular markers at the cellular level in esophageal cancer and gastric cancer, so as to provide more accurate prediction and detection. Diagnostic information.

SUMMARY OF THE INVENTION

In view of the deficiencies of the prior art and the actual needs, the present invention provides a grading model for detecting the benign and malignant degree of esophageal tumors and/or gastric tumors and its application. The detection device and model are used at the cellular and tissue level The changes of imprinted (trace) genes of esophageal tumors and/or gastric tumors can be visually observed at an early stage to judge the benign and malignant degree of esophageal tumors and/or gastric tumors.

To achieve the above object, the present invention adopts the following technical solutions:

In a first aspect, the present invention provides an imprinted gene grading model for esophageal tumors and/or gastric tumors, the model calculates the expression of missing imprinted genes, the abnormal expression of imprinted gene copy number and the total expression of imprinted genes. Changes in esophageal and/or gastric tumors grade the expression status of imprinted genes;

Wherein, the imprinted gene is any one or a combination of at least two of Z1, Z11 or Z16, the imprinted gene Z1 is Gnas, the imprinted gene Z11 is Grb10, and the imprinted gene Z16 is Snrpn/Snurf.

In the present invention, the inventors found that by calculating the expression of missing imprinted genes and the abnormal expression of imprinted gene copies in esophageal tumors and/or gastric tumors of any one of Z1, Z11 and Z16, the sensitivity of the diagnosis of esophageal cancer It can reach more than 90.4%, and the diagnostic sensitivity of gastric cancer can reach more than 67.6%.

According to the present invention, if only one imprinted gene is detected in the preliminary detection, any one of Z1, Z11 and Z16 can be detected. The most preferred imprinted gene for detecting esophageal cancer is Z1, and the most preferred imprinting gene for detecting gastric cancer is Z11.

In the present invention, the inventors found that the detection sensitivity of a single Z1 imprinted gene for esophageal cancer can reach 96.3%, and the detection of a single Z11 imprinted gene can reach 92.5% for esophageal cancer. Imprinted genes, the diagnostic sensitivity of esophageal cancer can reach 90.4%;

In the present invention, the inventor found that the detection sensitivity of a single Z1 imprinted gene for gastric cancer can reach 80.6%, and the detection of a single Z11 imprinted gene can reach 88.9% for gastric cancer, and a single Z16 imprinted gene can be detected. , the diagnostic sensitivity of gastric cancer can reach 67.6%.

According to the present invention, if a combination of two imprinted genes of an imprinted gene is detected, the combination can be any two of Z1, Z11 and Z16, for example, the combination can be a combination of Z1 and Z11, a combination of Z1 and Z16 or A combination of Z11 and Z16.

In the present invention, the inventors found that the sensitivity can be further improved by calculating the missing expression levels of the imprinted genes and the abnormal expression levels of the imprinted gene copy numbers of two or more imprinted genes. The diagnostic sensitivity of esophageal cancer can reach more than 98.1%. When the combination of Z1 and Z16 is detected, the diagnostic sensitivity of esophageal cancer can reach 98.1%. When the combination of Z1 and Z11 is detected, the diagnostic sensitivity of esophageal cancer can reach 99.2 %, when the combination of Z11 and Z16 is detected, the diagnostic sensitivity for esophageal cancer can reach 99.2%;

In the present invention, the inventors found that the sensitivity can be further improved by calculating the missing expression levels of the imprinted genes and the abnormal expression levels of the imprinted gene copy numbers of two or more imprinted genes. The diagnostic sensitivity of gastric cancer can reach more than 94.1%. When the combination of Z1 and Z16 is detected, the diagnostic sensitivity of gastric cancer can reach 97.1%. When the combination of Z1 and Z11 is detected, the diagnostic sensitivity of gastric cancer can reach 97.2%. When the combination of Z11 and Z16, the diagnostic sensitivity of gastric cancer can reach 94.1%.

According to the present invention, the imprinted gene further includes any one or a combination of at least two of Z3, Z4, Z5, Z6, Z8, Z10 or Z13; wherein, the imprinted gene Z3 is Peg10, and the imprinted gene Z4 is For Igf2r, the imprinted gene Z5 is Mest, the imprinted gene Z6 is Plagl1, the imprinted gene Z8 is Dcn, the imprinted gene Z10 is Gatm, and the imprinted gene Z13 is Sgce.

In the present invention, the inventors found that adding Z3, Z4, Z5, Z6, Z8, Z10, Z13 genes for joint diagnosis on the basis of Z1, Z11 and Z16 gene detection not only helps to increase the detection accuracy, but also Adding other probes to assist diagnosis can further avoid the occurrence of false positives, and can further improve the detection accuracy, so that accurate grading and judgment of all esophageal and gastric tumor samples can be achieved.

According to the present invention, the method for calculating imprinted genes by the model is: calculating the combination of imprinted genes, the imprinted genes being the combination of ten genes of Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 .

In the present invention, the deletion of the imprinted gene means that after the cells are stained with hematoxylin, there are two red/brown marks in the nucleus, and the abnormal copy number of the imprinted gene means that after the cells are stained with hematoxylin, there are more than two red/brown marks in the nucleus of the cell. Marked in brown, the copy number abnormality is due to abnormal gene duplication by cancer cells, resulting in a condition where this gene is expressed as a triploid or even higher polyploidy.

In the present invention, the hematoxylin-stained marker is selected from, but not limited to, red or brown, and staining with other colors can also be used for the calculation of the total expression of imprinted genes, the expression of missing imprinted genes, and the abnormal expression of imprinted gene copy numbers. .

In the present invention, the imprinted gene and the imprinted gene are the same concept, represent the same meaning, and can be replaced.

Preferably, the formula for calculating the total expression of imprinted genes, the missing expression of imprinted genes and the abnormal expression of imprinted gene copy number is as follows:

Total expression = (b+c+d)/(a+b+c+d)×100%;

Total expression of normal imprinted genes=b/(b+c+d)×100%;

Imprinted gene deletion gene expression (LOI)=c/(b+c+d)×100%;

Gene expression with abnormal copy number of imprinted genes (CNV)=d/(b+c+d)×100%;

Wherein, the a is the cell nucleus where there is no marker in the nucleus after the cells are stained with hematoxylin, and the imprinted gene is not expressed; the b is the cell where there is a red/brown marker in the nucleus after the cell is stained with hematoxylin, and the imprinted gene is present. The cell nucleus; the c means that after the cells are stained with hematoxylin, there are two red/brown marks in the nucleus, and the nuclei with missing imprinted genes; the d means that after the cells are stained with hematoxylin, there are more than two red/brown marks in the nucleus , nuclei with abnormal copy number of imprinted genes.

In the present invention, the hematoxylin-stained marker is selected from, but not limited to, red or brown, and dyeing markers with other colors can also be used for the calculation of the missing expression of imprinted genes, the abnormal expression of imprinted gene copy number and the total expression of imprinted genes .

In the present invention, the probe is subjected to in situ hybridization, and Hemotoxy (hematoxylin) cell nucleus staining is used to amplify the signal, and under a 40× or 60× microscope, the presence of imprinted genes in each nucleus, the absence of imprinted genes or the abnormal copy number are determined. , by calculating the expression of missing imprinted genes, the abnormal expression of imprinted gene copy number and the total expression of imprinted genes to determine the benign and malignant degree of tumor in the sample; since the section is only 10 μm, about 20% of the nuclei seen under the microscope are For incomplete nuclei, that is to say, there is a possibility of partial false negatives.

According to the present invention, the missing expression amount of the imprinted gene, the abnormal expression amount of the imprinted gene copy number and the total expression amount of the imprinted gene are divided into five different levels.

According to the present invention, at least 1200 cells were counted in the most positively expressed region of the sample by each probe for the ten imprinted genes of Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 Five different grades were divided into five different levels for the missing expression of imprinted genes, the abnormal expression of imprinted gene copy number and the total expression of imprinted genes.

Preferably, the five different grades for the deletion of imprinted gene expression level, the abnormal expression level of imprinted gene copy number and the total expression level of Z1 are:

Grade 0: The missing expression level of the imprinted gene Z1 is less than 16%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z1 is less than 2.5%, or the total expression level of the imprinted gene Z1 is less than 35%. one or a combination of at least two;

Level I: The missing expression level of the imprinted gene Z1 is 16-20%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z1 is 2.5-4%, or the total expression level of the imprinted gene Z1 is 35% -40% of any one or a combination of at least two;

Level II: The missing expression level of the imprinted gene Z1 is 20-25%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z1 is 4-6.5%, or the total expression level of the imprinted gene Z1 is 40% -50% of any one or a combination of at least two;

Grade III: The missing expression level of the imprinted gene Z1 is 25-30%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z1 is 6.5-8%, or the total expression level of the imprinted gene Z1 is 50% - 65% of any one or a combination of at least two;

Grade IV: The missing expression of the imprinted gene Z1 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z1 is greater than 8%, or the total expression of the imprinted gene Z1 is greater than 65%. one or a combination of at least two.

Preferably, the five different grades for the deletion of imprinted gene expression, the abnormal expression of imprinted gene copy number and the total expression for Z3 and Z13 are:

Grade 0: The missing expression levels of the imprinted genes Z3 and Z13 are less than 10%, the abnormal expression levels of the imprinted gene copy numbers of the imprinted genes Z3 and Z13 are less than 1%, or the total expression levels of the imprinted genes Z3 and Z13 Any one or a combination of at least two of less than 25%;

Level I: The imprinted genes Z3 and Z13 have 10-15% missing expression levels, and the imprinted genes Z3 and Z13 have 1-2% abnormal expression levels of imprinted gene copy numbers or the imprinted genes Z3 and Z13 The total expression level is 25-30% of any one or a combination of at least two;

Level II: The imprinted genes Z3 and Z13 have 15-25% missing imprinted gene expression, the imprinted genes Z3 and Z13 have 2-3% abnormal expression of imprinted gene copy numbers or the imprinted genes Z3 and Z13 The total expression level is 30-45% of any one or a combination of at least two;

Grade III: The imprinted genes Z3 and Z13 have 25-30% missing expression levels, the imprinted genes Z3 and Z13 have 3-5% abnormal expression levels of imprinted gene copy numbers or the imprinted genes Z3 and Z13 The total expression level is 45-55% of any one or a combination of at least two;

Grade IV: The imprinted genes Z3 and Z13 have missing expression levels of more than 30%, the imprinted gene copy numbers of the imprinted genes Z3 and Z13 have abnormal expression levels of more than 5%, or the total expression levels of the imprinted genes Z3 and Z13 Any one or a combination of at least two of more than 55%;

In the present invention, the imprinted genes Z3 and Z13 are independent of each other in terms of the missing expression level of the imprinted gene, the abnormal expression level of the imprinted gene copy number and the total expression level.

Preferably, the five different grades for the imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level of Z4, Z5, Z6, Z10 and Z11 are:

Level 0: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have missing expression levels of less than 16%, and the imprinted gene copy numbers of the imprinted genes Z4, Z5, Z6, Z10 and Z11 have abnormal expression levels of less than 1.5% Or the total expression level of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is less than 30% of any one or a combination of at least two of them;

Level I: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have 16-20% missing expression levels, and the imprinted genes Z4, Z5, Z6, Z10 and Z11 have abnormally expressed imprinted gene copy numbers of 16-20%. 1.5-2.5% or the total expression level of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is 30-40% of any one or a combination of at least two;

Level II: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have 20-25% missing imprinted gene expression, and the imprinted genes Z4, Z5, Z6, Z10 and Z11 have abnormally expressed imprinted gene copy numbers of 20-25%. 2.5-4% or the total expression level of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is 40-50% of any one or a combination of at least two;

Grade III: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have 25-30% missing imprinted gene expression, and the imprinted gene Z4, Z5, Z6, Z10 and Z11 have abnormally expressed imprinted gene copy numbers of 25-30%. 4-6% or the total expression level of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is 50-65% of any one or a combination of at least two;

Level IV: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have a missing expression of more than 30%, and the imprinted genes Z4, Z5, Z6, Z10 and Z11 have an abnormal expression of imprinted gene copy number greater than 6% Or the total expression of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is greater than 65% of any one or a combination of at least two;

In the present invention, the imprinted genes Z4, Z5, Z6, Z10 and Z11 are independent of each other in terms of the missing expression levels of imprinted genes, the abnormal expression levels of imprinted gene copy numbers and the total expression levels.

Preferably, when used to detect esophageal tumors, the five different grades for Z16 are divided into five different levels of expression of imprinted gene deletion, abnormal expression of imprinted gene copy number and total expression:

Grade 0: Any of the missing expression of the imprinted gene Z16 is less than 10%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z16 is less than 1%, or the total expression of the imprinted gene Z16 is less than 25%. one or a combination of at least two;

Level I: The imprinted gene Z16 has 10-15% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 1-2%, or the imprinted gene Z16 has a total expression of 25% -30% of any one or a combination of at least two;

Level II: The imprinted gene Z16 has 15-25% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 2-4%, or the total expression of the imprinted gene Z16 is 30% -40% of any one or a combination of at least two;

Level III: The imprinted gene Z16 has 25-30% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 4-6%, or the total expression of the imprinted gene Z16 is 40% -55% of any one or a combination of at least two;

Grade IV: Any of the missing expression of the imprinted gene Z16 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z16 is greater than 6%, or the total expression of the imprinted gene Z16 is greater than 55%. one or a combination of at least two;

Preferably, when used to detect gastric tumors, the five different grades for Z16 are divided into five different levels of expression of imprinted gene deletion, abnormal expression of imprinted gene copy number and total expression:

Grade 0: Any of the missing expression level of the imprinted gene Z16 is less than 15%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z16 is less than 1.5%, or the total expression level of the imprinted gene Z16 is less than 30%. one or a combination of at least two;

Level I: The imprinted gene Z16 has a missing expression level of 15-20%, the imprinted gene copy number abnormal expression level of the imprinted gene Z16 is 1.5-4%, or the total expression level of the imprinted gene Z16 is 30% -40% of any one or a combination of at least two;

Level II: The imprinted gene Z16 has 20-25% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 4-7%, or the total expression of the imprinted gene Z16 is 40% -50% of any one or a combination of at least two;

Level III: The imprinted gene Z16 has 25-30% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 7-10%, or the total expression of the imprinted gene Z16 is 50% - 60% of any one or a combination of at least two;

Grade IV: Any of the missing expression of the imprinted gene Z16 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z16 is greater than 10%, or the total expression of the imprinted gene Z16 is greater than 60%. one or a combination of at least two.

Due to the specificity of the expression of the imprinted gene Z16 in gastric cancer, most of the grades III and IV of Z16 appear in the early and middle stages of gastric cancer, and 90% of advanced gastric cancer samples have low Z16 expression.

Preferably, when used for the detection of esophageal tumors, the five different grades for the Z8 imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level are:

Grade 0: The missing expression level of the imprinted gene Z8 is less than 16%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is less than 2.5%, or the total expression level of the imprinted gene Z8 is less than 35%. one or a combination of at least two;

Level I: The missing expression level of the imprinted gene Z8 is 16-20%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 2.5-4%, or the total expression level of the imprinted gene Z8 is 35% -40% of any one or a combination of at least two;

Level II: The missing expression level of the imprinted gene Z8 is 20-25%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 4-6.5%, or the total expression level of the imprinted gene Z8 is 40% -50% of any one or a combination of at least two;

Grade III: The missing expression level of the imprinted gene Z8 is 25-30%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 6.5-8%, or the total expression level of the imprinted gene Z8 is 50% - 65% of any one or a combination of at least two;

Grade IV: The missing expression of the imprinted gene Z8 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z8 is greater than 8%, or the total expression of the imprinted gene Z8 is greater than 65%. one or a combination of at least two;

Preferably, when used for the detection of gastric tumors, the five different grades for the Z8 imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level are:

Grade 0: The missing expression level of the imprinted gene Z8 is less than 10%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is less than 1%, or the total expression level of the imprinted gene Z8 is less than 25%. one or a combination of at least two;

Level I: The missing expression level of the imprinted gene Z8 is 10-15%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 1-2%, or the total expression level of the imprinted gene Z8 is 25% -30% of any one or a combination of at least two;

Level II: The missing expression level of the imprinted gene Z8 is 15-25%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 2-3%, or the total expression level of the imprinted gene Z8 is 30% -45% of any one or a combination of at least two;

Grade III: The missing expression level of the imprinted gene Z8 is 25-30%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 3-5%, or the total expression level of the imprinted gene Z8 is 45% -55% of any one or a combination of at least two;

Grade IV: The missing expression of the imprinted gene Z8 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z8 is greater than 5%, or the total expression of the imprinted gene Z8 is greater than 55%. one or a combination of at least two.

In the present invention, the inventors found that the imprinted genes Z8 and Z16 are special, and their grading of the missing expression of imprinted genes, the abnormal expression of imprinted gene copy number and the total expression in esophageal tumors or gastric tumors are different. The imprinted genes Z8 and Z16 were graded separately for esophageal or gastric tumors.

In a second aspect, the present invention provides a device for detecting the benign and malignant degree of esophageal tumor and/or gastric tumor, comprising the following units:

(1) Sampling unit: obtain the sample to be tested;

(2) Probe design unit: design specific primers according to the imprinted gene sequence;

(3) detection unit: perform in situ hybridization with the probe of step (2) and the sample to be tested;

(4) Analysis unit: microscope imaging to analyze the expression of imprinted genes;

Wherein, the analysis unit calculates the total expression of imprinted genes, the expression of missing imprinted genes, and the abnormal expression of imprinted gene copy numbers, and uses the imprinted gene grading model described in the first aspect, so as to calculate the missing expression of imprinted genes, the expression of imprinted genes, and the abnormal expression of imprinted gene copies. The level of abnormal expression of copy number and total expression of imprinted genes was used to judge the benign and malignant degree of esophageal tumor and/or gastric tumor.

In the present invention, the deletion of the imprinted gene means that after the cells are stained with hematoxylin, there are two red/brown marked nuclei in the nucleus, and the abnormal copy number of the imprinted gene means that after the cells are stained with hematoxylin, there are more than two nuclei in the nucleus Red/brown marked nuclei, the copy number abnormality is due to abnormal gene duplication by cancer cells, resulting in a situation where this gene is expressed as a triploid or even higher polyploidy.

In the present invention, the hematoxylin-stained marker is selected from, but not limited to, red or brown, and other colors for staining markers can also be used to calculate the total expression of imprinted genes, the missing expression of imprinted genes, and the abnormal expression of imprinted gene copy numbers. .

The detection system of the present invention is used to visually observe the changes of imprinted (trace) genes of esophagus and gastric tumors at the cell and tissue level in the early stage, so as to judge the benign and malignant degree and malignant degree of the tumor, and provide the most favorable conditions for patients with early esophageal and gastric tumors. treatment opportunities.

According to the present invention, the sample to be tested in step (1) is derived from human tissue and/or cells.

In the present invention, the sample to be tested is feasible as long as the RNA is fixed in time, and those skilled in the art can select it as needed, which is not particularly limited here. The sample to be tested in the present invention includes paraffin sections of tissues , any one or a combination of at least two of esophageal mucosal exfoliated cells or gastroscopic biopsy samples.

The specific operation steps of the paraffin section of the tissue are to obtain a human tumor tissue sample, fix it with 10% neutral formalin in time, embed it in paraffin, cut it into 10 μm thick, and use a positively charged glass slide to make a tissue slide; because It is only 10 μm thick, so some of the nuclei seen under the microscope are incomplete, so there will be some false negative gene deletions.

The specific operation steps of the esophageal mucosal exfoliated cells are to obtain the mucosal exfoliated cells through the esophagus pull-net inspection, and fix them with 10% neutral formalin in time.

The specific operation steps of the gastroscopic biopsy sample are to obtain the human cells of the esophagus or stomach under the gastroscope, and fix it with 10% neutral formalin in time.

In the present invention, since the principle of obtaining esophageal mucosal exfoliated cells is similar to that of gastroscopic biopsy samples, the operation mode of the sample detection is similar to that of gastroscopic biopsy samples, and the results are also similar.

In the present invention, the esophageal mucosal exfoliated cells and gastroscopic biopsy are less harmful to the patient and the sampling process is simple. Compared with the circulation characteristics of blood, the esophageal mucosal exfoliated cells and gastroscopic biopsy can also be located, and the esophageal mucosal exfoliated cells and gastroscopic biopsy are used as experiments. Samples have their special advantages.

Preferably, the sample to be tested is esophageal mucosal exfoliated cells and/or a gastroscopic biopsy sample.

Preferably, the imprinted genes are Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16, the imprinted gene Z1 is Gnas, the imprinted gene Z3 is Peg10, and the imprinted gene Z4 is Igf2r, the imprinted gene Z5 is Mest, the imprinted gene Z6 is Plagl1, the imprinted gene Z8 is Dcn, the imprinted gene Z10 is Gatm, the imprinted gene Z11 is Grb10, the imprinted gene Z13 is Sgce, The imprinted gene Z16 is Snrpn/Snurf.

In the present invention, the imprinted genes Z1 (Gnas), Z3 (Peg10), Z4 (Igf2r), Z5 (Mest), Z6 (Plagl1), Z8 (Dcn), Z10 (Gatm), Z11 (Grb10), Z13 ( Sgce), Z16 (Snrpn/Snurf) are expressed in different degrees in normal tumor cells, and the expression and imprinting state will change significantly when malignant lesions occur.

In the present invention, the designed probes are based on imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16, namely Gnas, Peg10, Igf2r, Mest, Plagl1, Dcn, Gatm, Grb10, Designed by Sgce and Snrpn/Snurf, a sequence is selected as a probe in the endoroton of each gene, and the specific probe is designed by Advanced CellDiagnostics.

Preferably, the in situ hybridization adopts RNAscope in situ hybridization method.

Preferably, the RNAscope in situ hybridization method uses a single-channel or multi-channel coloring kit or a single-channel or multi-channel fluorescence kit, preferably a single-channel red/brown coloring kit or a multi-channel fluorescence kit .

In the present invention, the multi-channel color-forming kit or multi-channel fluorescence kit includes two-channel or more than two-channel color-forming kits or fluorescence kits, and the two-channel color-forming kit or multi-channel fluorescence reagent Cassettes can use two imprinted gene probes or the combined expression of imprinted and other genes or even the combined expression of multiple imprinted and non-imprinted genes.

According to the present invention, the formulas for calculating the total expression of imprinted genes, the missing expression of imprinted genes and the abnormal expression of imprinted gene copy number in the model are as follows:

Total expression = (b+c+d)/(a+b+c+d)×100%;

Total expression of normal imprinted genes=b/(b+c+d)×100%;

Imprinted gene deletion gene expression (LOI)=c/(b+c+d)×100%;

Gene expression with abnormal copy number of imprinted genes (CNV)=d/(b+c+d)×100%;

Wherein, the a is the cell nucleus where there is no marker in the nucleus after the cells are stained with hematoxylin, and the imprinted gene is not expressed; the b is the cell where there is a red/brown marker in the nucleus after the cell is stained with hematoxylin, and the imprinted gene is present. The cell nucleus; the c means that after the cells are stained with hematoxylin, there are two red/brown marks in the nucleus, and the nuclei with missing imprinted genes; the d means that after the cells are stained with hematoxylin, there are more than two red/brown marks in the nucleus , nuclei with abnormal copy number of imprinted genes.

In the present invention, the hematoxylin-stained marker is selected from, but not limited to, red or brown, and staining with other colors can also be used for the calculation of the total expression of imprinted genes, the expression of missing imprinted genes, and the abnormal expression of imprinted gene copy numbers. .

In the present invention, the probe is subjected to in situ hybridization, and Hemotoxy (hematoxylin) cell nucleus staining is used to amplify the signal, and under a 40× or 60× microscope, the presence of imprinted genes in each nucleus, the absence of imprinted genes or the abnormal copy number are determined. , by calculating the total expression of imprinted genes, the expression of missing imprinted genes, and the gene expression of abnormal imprinted gene copy numbers to determine the benign and malignant degree of the tumor in the sample. Since the section is only 10 microns, about 20% of the nuclei seen under the microscope are incomplete nuclei, which means there is a possibility of some false negatives.

According to the present invention, the missing expression level of the imprinted gene, the abnormal expression level of the imprinted gene copy number and the total expression level are divided into five different levels.

The five different grades are counts of at least 1200 cells in the region of the sample where each probe expresses the most positive, ten for Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 The missing expression of imprinted genes, the abnormal expression of imprinted gene copy number and the total expression of imprinted genes were divided.

Preferably, the five different grades for the deletion of imprinted gene expression level, the abnormal expression level of imprinted gene copy number and the total expression level of Z1 are:

Grade 0: The missing expression level of the imprinted gene Z1 is less than 16%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z1 is less than 2.5%, or the total expression level of the imprinted gene Z1 is less than 35%. one or a combination of at least two;

Level I: The missing expression level of the imprinted gene Z1 is 16-20%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z1 is 2.5-4%, or the total expression level of the imprinted gene Z1 is 35% -40% of any one or a combination of at least two;

Level II: The missing expression level of the imprinted gene Z1 is 20-25%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z1 is 4-6.5%, or the total expression level of the imprinted gene Z1 is 40% -50% of any one or a combination of at least two;

Grade III: The missing expression level of the imprinted gene Z1 is 25-30%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z1 is 6.5-8%, or the total expression level of the imprinted gene Z1 is 50% - 65% of any one or a combination of at least two;

Grade IV: The missing expression of the imprinted gene Z1 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z1 is greater than 8%, or the total expression of the imprinted gene Z1 is greater than 65%. one or a combination of at least two.

Preferably, the five different grades for the deletion of imprinted gene expression, the abnormal expression of imprinted gene copy number and the total expression for Z3 and Z13 are:

Grade 0: The missing expression levels of the imprinted genes Z3 and Z13 are less than 10%, the abnormal expression levels of the imprinted gene copy numbers of the imprinted genes Z3 and Z13 are less than 1%, or the total expression levels of the imprinted genes Z3 and Z13 Any one or a combination of at least two of less than 25%;

Level I: The imprinted genes Z3 and Z13 have 10-15% missing expression levels, and the imprinted genes Z3 and Z13 have 1-2% abnormal expression levels of imprinted gene copy numbers or the imprinted genes Z3 and Z13 The total expression level is 25-30% of any one or a combination of at least two;

Level II: The imprinted genes Z3 and Z13 have 15-25% missing imprinted gene expression, the imprinted genes Z3 and Z13 have 2-3% abnormal expression of imprinted gene copy numbers or the imprinted genes Z3 and Z13 The total expression level is 30-45% of any one or a combination of at least two;

Grade III: The imprinted genes Z3 and Z13 have 25-30% missing expression levels, the imprinted genes Z3 and Z13 have 3-5% abnormal expression levels of imprinted gene copy numbers or the imprinted genes Z3 and Z13 The total expression level is 45-55% of any one or a combination of at least two;

Grade IV: The imprinted genes Z3 and Z13 have missing expression levels of more than 30%, the imprinted gene copy numbers of the imprinted genes Z3 and Z13 have abnormal expression levels of more than 5%, or the total expression levels of the imprinted genes Z3 and Z13 Any one or a combination of at least two of more than 55%;

In the present invention, the imprinted genes Z3 and Z13 are independent of each other in terms of the missing expression level of the imprinted gene, the abnormal expression level of the imprinted gene copy number and the total expression level.

Preferably, the five different grades for the imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level of Z4, Z5, Z6, Z10 and Z11 are:

Level 0: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have missing expression levels of less than 16%, and the imprinted gene copy numbers of the imprinted genes Z4, Z5, Z6, Z10 and Z11 have abnormal expression levels of less than 1.5% Or the total expression level of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is less than 30% of any one or a combination of at least two of them;

Level I: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have 16-20% missing expression levels, and the imprinted genes Z4, Z5, Z6, Z10 and Z11 have abnormally expressed imprinted gene copy numbers of 16-20%. 1.5-2.5% or the total expression level of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is 30-40% of any one or a combination of at least two;

Level II: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have 20-25% missing imprinted gene expression, and the imprinted genes Z4, Z5, Z6, Z10 and Z11 have abnormally expressed imprinted gene copy numbers of 20-25%. 2.5-4% or the total expression level of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is 40-50% of any one or a combination of at least two;

Grade III: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have 25-30% missing imprinted gene expression, and the imprinted gene Z4, Z5, Z6, Z10 and Z11 have abnormally expressed imprinted gene copy numbers of 25-30%. 4-6% or the total expression level of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is 50-65% of any one or a combination of at least two;

Level IV: The imprinted genes Z4, Z5, Z6, Z10 and Z11 have a missing expression of more than 30%, and the imprinted genes Z4, Z5, Z6, Z10 and Z11 have an abnormal expression of imprinted gene copy number greater than 6% Or the total expression of the imprinted genes Z4, Z5, Z6, Z10 and Z11 is greater than 65% of any one or a combination of at least two;

In the present invention, the imprinted genes Z4, Z5, Z6, Z10 and Z11 are independent of each other in terms of the missing expression levels of imprinted genes, the abnormal expression levels of imprinted gene copy numbers and the total expression levels.

Preferably, when used to detect esophageal tumors, the five different grades for Z16 are divided into five different levels of expression of imprinted gene deletion, abnormal expression of imprinted gene copy number and total expression:

Grade 0: Any of the missing expression of the imprinted gene Z16 is less than 10%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z16 is less than 1%, or the total expression of the imprinted gene Z16 is less than 25%. one or a combination of at least two;

Level I: The imprinted gene Z16 has 10-15% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 1-2%, or the imprinted gene Z16 has a total expression of 25% -30% of any one or a combination of at least two;

Level II: The imprinted gene Z16 has 15-25% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 2-4%, or the total expression of the imprinted gene Z16 is 30% -40% of any one or a combination of at least two;

Level III: The imprinted gene Z16 has 25-30% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 4-6%, or the total expression of the imprinted gene Z16 is 40% -55% of any one or a combination of at least two;

Grade IV: Any of the missing expression of the imprinted gene Z16 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z16 is greater than 6%, or the total expression of the imprinted gene Z16 is greater than 55%. one or a combination of at least two.

Preferably, when used to detect gastric tumors, the five different grades for Z16 are divided into five different levels of expression of imprinted gene deletion, abnormal expression of imprinted gene copy number and total expression:

Grade 0: Any of the missing expression level of the imprinted gene Z16 is less than 15%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z16 is less than 1.5%, or the total expression level of the imprinted gene Z16 is less than 30%. one or a combination of at least two;

Level I: The imprinted gene Z16 has a missing expression level of 15-20%, the imprinted gene copy number abnormal expression level of the imprinted gene Z16 is 1.5-4%, or the total expression level of the imprinted gene Z16 is 30% -40% of any one or a combination of at least two;

Level II: The imprinted gene Z16 has 20-25% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 4-7%, or the total expression of the imprinted gene Z16 is 40% -50% of any one or a combination of at least two;

Level III: The imprinted gene Z16 has 25-30% missing imprinted gene expression, the imprinted gene Z16 has an abnormally expressed imprinted gene copy number of 7-10%, or the total expression of the imprinted gene Z16 is 50% - 60% of any one or a combination of at least two;

Grade IV: Any of the missing expression of the imprinted gene Z16 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z16 is greater than 10%, or the total expression of the imprinted gene Z16 is greater than 60%. one or a combination of at least two.

Due to the specificity of the expression of the imprinted gene Z16 in gastric cancer, most of the grades III and IV of Z16 appear in the early and middle stages of gastric cancer, and 90% of advanced gastric cancer samples have low Z16 expression.

Preferably, when used for the detection of esophageal tumors, the five different grades for the Z8 imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level are:

Grade 0: The missing expression level of the imprinted gene Z8 is less than 16%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is less than 2.5%, or the total expression level of the imprinted gene Z8 is less than 35%. one or a combination of at least two;

Level I: The missing expression level of the imprinted gene Z8 is 16-20%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 2.5-4%, or the total expression level of the imprinted gene Z8 is 35% -40% of any one or a combination of at least two;

Level II: The missing expression level of the imprinted gene Z8 is 20-25%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 4-6.5%, or the total expression level of the imprinted gene Z8 is 40% -50% of any one or a combination of at least two;

Grade III: The missing expression level of the imprinted gene Z8 is 25-30%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 6.5-8%, or the total expression level of the imprinted gene Z8 is 50% - 65% of any one or a combination of at least two;

Grade IV: The missing expression of the imprinted gene Z8 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z8 is greater than 8%, or the total expression of the imprinted gene Z8 is greater than 65%. one or a combination of at least two;

Preferably, when used for the detection of gastric tumors, the five different grades for the Z8 imprinted gene deletion expression level, the imprinted gene copy number abnormal expression level and the total expression level are:

Grade 0: The missing expression level of the imprinted gene Z8 is less than 10%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is less than 1%, or the total expression level of the imprinted gene Z8 is less than 25%. one or a combination of at least two;

Level I: The missing expression level of the imprinted gene Z8 is 10-15%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 1-2%, or the total expression level of the imprinted gene Z8 is 25% -30% of any one or a combination of at least two;

Level II: The missing expression level of the imprinted gene Z8 is 15-25%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 2-3%, or the total expression level of the imprinted gene Z8 is 30% -45% of any one or a combination of at least two;

Grade III: The missing expression level of the imprinted gene Z8 is 25-30%, the abnormal expression level of the imprinted gene copy number of the imprinted gene Z8 is 3-5%, or the total expression level of the imprinted gene Z8 is 45% -55% of any one or a combination of at least two;

Grade IV: The missing expression of the imprinted gene Z8 is greater than 30%, the abnormal expression of the imprinted gene copy number of the imprinted gene Z8 is greater than 5%, or the total expression of the imprinted gene Z8 is greater than 55%. one or a combination of at least two.

In the present invention, the inventors found that the imprinted genes Z8 and Z16 are special, and the grading of the missing imprinted gene expression, the abnormal expression of imprinted gene copy number and the total expression of these two genes in esophageal tumors or gastric tumors are different. , so the imprinted genes Z8 and Z16 were graded separately according to esophageal tumors or gastric tumors.

In the present invention, the imprinted gene Z16 is uniformly expressed in the interstitial and epithelial cells of gastric tumor tissue, but is significantly different in early gastric cancer before clinical stage II and intermediate and advanced gastric cancer in clinical stage III-IV.

Preferably, the judging the benign and malignant degree of esophageal tumor is divided into benign tumor, esophageal cancer potential, early esophageal cancer, intermediate esophageal cancer and advanced esophageal cancer.

Preferably, the result of judging the benign and malignant degree of the esophagus is that the missing expression levels of imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 and the abnormal expression levels of imprinted gene copy numbers are both Less than grade I, the imprinted gene deletion expression level of no more than one imprinted gene in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade I and the imprinted genes Z1, Z3, Z4 , Z5, Z6, Z8, Z10, Z11, Z13 and Z16, the abnormal expression of imprinted gene copy number of no more than one imprinted gene is any one of grade I, then it is a benign tumor;

Preferably, the result of judging the benign and malignant degree of the esophagus is that the imprinted gene deletion expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is 1 At least two imprinted genes of Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13, and Z16 have abnormal expression levels of imprinted gene copy number at level I or imprinted genes Z1, Z3, Z4, Z5 , Z6, Z8, Z10, Z11, Z13 and Z16 with no more than 1 imprinted gene missing expression level II and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 If the abnormal expression of imprinted gene copy number of no more than 1 imprinted gene is any one of grade II, it is esophageal cancer potential;

Preferably, the result of judging the benign and malignant degree of the esophagus is that the expression level of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is II. Grade II, the imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is abnormal expression level II or imprinted genes Z1, Z3, Z4, Z5 , Z6, Z8, Z10, Z11, Z13 and Z16 with no more than 1 imprinted gene missing expression level III and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 If the abnormal expression of imprinted gene copy number of no more than 1 imprinted gene is any one of grade III, it is early esophageal cancer;

Preferably, the result of judging the benign and malignant degree of the esophagus is that the expression level of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is III. Grade III, the imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is abnormal expression level III or imprinted genes Z1, Z3, Z4, Z5 , Z6, Z8, Z10, Z11, Z13 and Z16 with no more than 1 imprinted gene missing expression level IV and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 If the abnormal expression of imprinted gene copy number of no more than 1 imprinted gene is any one of grade IV, it is mid-stage esophageal cancer;

Preferably, the result of judging the benign and malignant degree of the esophagus is that the imprinted gene deletion expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade IV Or the abnormal expression of imprinted gene copy number of at least 2 imprinted genes in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade IV, which is advanced esophageal cancer.

Preferably, the judging the benign and malignant degree of gastric tumor is divided into benign gastric tumor, gastric cancer potential, early gastric cancer, intermediate gastric cancer and advanced gastric cancer.

Preferably, the result of judging the benign and malignant degree of gastric tumor is the expression level of imprinted gene deletion and the abnormal expression level of imprinted gene copy number of imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 All are less than grade I, and the expression level of imprinted genes with no more than one imprinted gene in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade I, and the imprinted genes Z1, Z3, The abnormal expression of imprinted gene copy number of no more than 1 imprinted gene in Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is any one of grade I, which is a benign tumor;

Preferably, the result of judging the benign and malignant degree of gastric tumor is that the expression level of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level I deletion , the abnormal expression of imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 was grade I, and the imprinted genes Z1, Z3, Z4, Z5, Z6, Imprinted gene deletion expression level of no more than 1 imprinted gene in Z8, Z10, Z11 and Z13 is level I and no more than 1 of the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 The abnormal expression level of imprinted gene copy number of imprinted genes is level I and the imprinted gene Z16 has missing expression level or abnormal expression level of imprinted gene copy number is level I or above, or imprinted genes Z1, Z3, Z4, Z5, Z6, Imprinted gene deletion expression level II for no more than 1 imprinted gene in Z8, Z10, Z11 and Z13 and no more than 1 imprint in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 If the abnormal expression level of imprinted gene copy number of the gene is any one of grade II, it is gastric cancer potential;

Preferably, the result of judging the benign and malignant degree of gastric tumor is that the expression level of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level II. , the imprinted gene copy number abnormal expression level of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level II or the imprinted genes Z1, Z3, Z4, Z5, Z6 , Z8, Z10, Z11 and Z13 no more than 1 imprinted gene expression level III imprinted gene deletion and no more than 1 imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 If the abnormal expression of imprinted gene copy number of imprinted genes is any one of grade III, it is early gastric cancer;

Preferably, the result of judging the benign and malignant degree of gastric tumor is that the expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level III. , the abnormal expression of imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level III or imprinted genes Z1, Z3, Z4, Z5, Z6, Imprinted gene deletion expression level of no more than 1 imprinted gene in Z8, Z10, Z11 and Z13 is grade IV and no more than 1 imprinted in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 If the abnormal expression level of imprinted gene copy number of the gene is any one of grade IV, it is mid-stage gastric cancer;

Preferably, the result of judging the benign and malignant degree of gastric tumor is that the imprinted gene deletion expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is grade IV or If the abnormal expression of imprinted gene copy number of at least 2 imprinted genes in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is grade IV, it is considered to be advanced gastric cancer.

In a third aspect, the present invention provides an imprinted gene grading model according to the first aspect and/or the device according to the second aspect for preparing a drug or kit for the detection and/or treatment of esophageal tumors and gastric tumors.

Preferably, judging the benign and malignant degree of esophageal tumor is divided into benign tumor, esophageal cancer potential, early esophageal cancer, intermediate esophageal cancer and advanced esophageal cancer.

Preferably, the result of judging the benign and malignant degree of esophageal tumor is the expression level of imprinted gene deletion and the abnormal expression level of imprinted gene copy number of imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 All are less than grade I, and the expression level of imprinted genes with no more than one imprinted gene in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade I, and the imprinted genes Z1, Z3, The abnormal expression of imprinted gene copy number of no more than 1 imprinted gene in Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is any one of grade I, which is a benign tumor;

Preferably, the result of judging the benign and malignant degree of esophageal tumor is that the expression levels of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 are missing Grade I, the abnormal expression of imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is Grade I or imprinted genes Z1, Z3, Z4, Imprinted gene deletion expression level of no more than 1 imprinted gene in Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is level II and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and If the abnormal expression of imprinted gene copy number of no more than one imprinted gene in Z16 is any one of grade II, it is esophageal cancer potential;

Preferably, the result of judging the benign and malignant degree of esophageal tumor is that the expression levels of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 are missing Grade II, the abnormal expression of imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade II or imprinted genes Z1, Z3, Z4, Imprinted gene deletion expression level of no more than 1 imprinted gene in Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is level III and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and If the abnormal expression of imprinted gene copy number of no more than one imprinted gene in Z16 is any one of grade III, it is early esophageal cancer;

Preferably, the result of judging the benign and malignant degree of esophageal tumor is that the expression levels of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 are missing Grade III, the abnormal expression of imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade III or imprinted genes Z1, Z3, Z4, Imprinted gene deletion expression level of no more than 1 imprinted gene in Z5, Z6, Z8, Z10, Z11, Z13 and Z16 was grade IV and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and If the abnormal expression of imprinted gene copy number of no more than one imprinted gene in Z16 is any one of grade IV, it is intermediate-stage esophageal cancer;

Preferably, the result of judging the benign and malignant degree of esophageal tumor is that the expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is IV. Advanced esophageal cancer was defined as grade IV or abnormal expression of imprinted gene copy number of at least 2 imprinted genes in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16.

Preferably, the judging the benign and malignant degree of gastric tumor is divided into benign gastric tumor, gastric cancer potential, early gastric cancer, intermediate gastric cancer and advanced gastric cancer.

Preferably, the result of judging the benign and malignant degree of gastric tumor is the expression level of imprinted gene deletion and the abnormal expression level of imprinted gene copy number of imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 All are less than grade I, and the expression level of imprinted genes with no more than one imprinted gene in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade I, and the imprinted genes Z1, Z3, The abnormal expression of imprinted gene copy number of no more than 1 imprinted gene in Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is any one of grade I, which is a benign tumor;

Preferably, the result of judging the benign and malignant degree of gastric tumor is that the expression level of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level I deletion , the abnormal expression of imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 was grade I, and the imprinted genes Z1, Z3, Z4, Z5, Z6, Imprinted gene deletion expression level of no more than 1 imprinted gene in Z8, Z10, Z11 and Z13 is level I and no more than 1 of the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 The abnormal expression level of imprinted gene copy number of imprinted genes is level I and the imprinted gene Z16 has missing expression level or abnormal expression level of imprinted gene copy number is level I or above, or imprinted genes Z1, Z3, Z4, Z5, Z6, Imprinted gene deletion expression level II for no more than 1 imprinted gene in Z8, Z10, Z11 and Z13 and no more than 1 imprint in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 If the abnormal expression level of imprinted gene copy number of the gene is any one of grade II, it is gastric cancer potential;

Preferably, the result of judging the benign and malignant degree of gastric tumor is that the expression level of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level II. , the imprinted gene copy number abnormal expression level of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level II or the imprinted genes Z1, Z3, Z4, Z5, Z6 , Z8, Z10, Z11 and Z13 no more than 1 imprinted gene expression level III imprinted gene deletion and no more than 1 imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 If the abnormal expression of imprinted gene copy number of imprinted genes is any one of grade III, it is early gastric cancer;

Preferably, the result of judging the benign and malignant degree of gastric tumor is that the expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level III. , the abnormal expression of imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level III or imprinted genes Z1, Z3, Z4, Z5, Z6, Imprinted gene deletion expression level of no more than 1 imprinted gene in Z8, Z10, Z11 and Z13 is grade IV and no more than 1 imprinted in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 If the abnormal expression level of imprinted gene copy number of the gene is any one of grade IV, it is mid-stage gastric cancer;

Preferably, the result of judging the benign and malignant degree of gastric tumor is that the imprinted gene deletion expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is grade IV or If the abnormal expression of imprinted gene copy number of at least 2 imprinted genes in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is grade IV, it is considered to be advanced gastric cancer.

In a fourth aspect, the present invention provides a marker for the diagnosis of advanced gastric cancer, the markers are imprinted gene Z1 and imprinted gene Z11;

Wherein, the imprinted gene Z1 is Gnas, and the imprinted gene Z11 is Grb10;

According to the present invention, the advanced gastric cancer is gastric cancer whose TNM grade is greater than T2N0M0.

In the present invention, the imprinted gene Z16 is uniformly expressed in the interstitial and epithelial cells of gastric tumor tissue, but is significantly different in early gastric cancer before clinical stage II and intermediate and advanced gastric cancer in clinical stage III-IV. In early gastric cancer, especially in the stage of intraepithelial neoplasia, 75% of the samples have imprinted deletion, copy number abnormality and total expression level of the imprinted gene Z16 reaching grade I or above. The imprinting deletion, copy number abnormality and total expression level of the imprinted gene Z16 were grade 0. Therefore, the imprinted gene Z16 combined with the imprinted genes Z1 and Z11 is a good diagnostic marker for the diagnosis of early gastric cancer, especially the transition from low-grade intraepithelial neoplasia to gastric cancer, but the imprinted gene Z16 is useful in advanced gastric cancer. Not much, only the Z1 and Z11 are needed to make a diagnosis.

Compared with the prior art, the present invention has the following beneficial effects:

(1) The detection model and system of the present invention express the performance of imprinting deletion on samples of patients with esophagus and gastric tumors in an intuitive way, and through the method of in situ labeling of imprinted genes, objective, intuitive, early and accurate Changes in imprinted (trace) genes can be detected, and a quantitative model can be provided, making a great contribution to the diagnosis of esophageal and gastric tumors;

(2) The detection device of the present invention can judge the benign and malignant degree of esophagus and gastric tumors and the invasion range of cancer cells through gastroscopic biopsy samples before the operation of patients with esophageal and gastric tumors, thereby providing a basis for surgery and precise treatment. Revolutionary breakthrough in the diagnosis of esophageal and gastric tumors in the molecular field;

(3) The present invention can accurately judge the grades of esophagus and gastric tumors, and clearly grade the malignant degree of early lesions of esophageal and gastric tumors, that is, intraepithelial neoplasia, through the combined detection of imprinted genes, especially the early and middle stage. Accurately distinguishing advanced gastric cancer greatly improves the early and definite diagnosis of esophageal cancer and gastric cancer, especially for early screening and follow-up after cancer surgery, especially for the follow-up of patients suspected of recurrence, which can buy time and save patients. make a significant contribution to life;

(4) The detection method of the present invention is different from the immunohistochemical method, and reduces false positives and other negative effects. Not only that, the gene silencing, deletion and rearrangement caused by the found deletion sites of imprinted genes related to esophagus and gastric tumors. Targeted drugs or technical methods can be used to guide later treatment and medication.

Description of drawings

Fig. 1 is a pathological section of gastric cancer with hematoxylin-stained nuclei of the present invention, wherein the a is that after the cells are stained with hematoxylin, there is no marker in the nucleus, and the imprinted genes are not expressed; There is one red/brown marker, and the imprinted gene exists; the c means that after the cells are stained with hematoxylin, there are two red/brown marks in the nucleus, and the imprinted gene is missing; the d is the cell nucleus after hematoxylin staining. Abnormal copy number of imprinted genes in more than two red/brown markers;

Figure 2(a) is the expression status of 10 genes in the pathological section of grade 0 gastric tumor, Figure 2(b) is the expression status of 10 genes in the pathological section of grade I gastric cancer, and Figure 2(c) is the grade II gastric cancer The expression status of 10 genes in the pathological section of the 1st grade gastric cancer, Figure 2(d) is the expression status of 10 genes in the pathological section of grade III gastric cancer, and Figure 2(e) is the expression status of 10 genes in the pathological section of grade IV gastric cancer ;

Figure 3(a) is the intensity of imprinting deletion of imprinted genes Z1, Z11 and Z16 in gastric cancer, Figure 3(b) is the intensity of imprinted genes Z1, Z11 and Z16 in gastric cancer copy number abnormality, Figure 3(c) is the imprinting The intensity of the total expression of genes Z1, Z11 and Z16 in gastric cancer, Figure 3(d) is the intensity of imprinted deletion of the imprinted genes Z3, Z4, Z5, Z6, Z8, Z10 and Z13 in gastric cancer, Figure 3(e) is The intensity of copy number abnormalities of imprinted genes Z3, Z4, Z5, Z6, Z8, Z10 and Z13 in gastric cancer, Figure 3(f) is the total expression of imprinted genes Z3, Z4, Z5, Z6, Z8, Z10 and Z13 in gastric cancer where LOI is the expression level of missing imprinted genes, CNV is the gene expression level with abnormal copy number of imprinted genes, and TE is the total expression level of imprinted genes;

Figure 4(a) is the intensity of imprinted deletion, copy number abnormality and total expression of imprinted gene Z1, Figure 4(b) is the intensity of imprinted deletion, copy number abnormality and total expression of imprinted gene Z11, and Figure 4(c) is The intensity of imprinting deletion, copy number abnormality and total expression of imprinted gene Z16, Fig. 4(d) is the intensity of imprinting deletion, copy number abnormality and total expression of imprinted gene Z3, Fig. 4(e) is imprinted gene Z4 imprinting deletion, The intensity of copy number abnormality and total expression, Figure 4(f) is the intensity of imprinting deletion, copy number abnormality and total expression of imprinted gene Z5, and Figure 4(g) is the imprinting deletion, copy number abnormality and total expression of imprinted gene Z6 Figure 4(h) is the intensity of imprinted deletion, copy number abnormality and total expression of imprinted gene Z8, Figure 4(i) is the intensity of imprinted deletion, copy number abnormality and total expression of imprinted gene Z10, Figure 4 (j) is the intensity of imprinting deletion, copy number abnormality and total expression level of imprinted gene Z13, where LOI is the expression level of imprinted gene deletion gene, CNV is the gene expression level of imprinted gene copy number abnormality, and TE is the total expression level of imprinted gene ;

Figure 5(a) shows the distribution range and grading standard of imprinting deletion and copy number abnormality in 37 cases of gastric tumor pathological sections of imprinted gene Z1, and Figure 5(b) shows the application of imprinted gene Z11 in 37 cases of gastric tumor pathological sections , the distribution range and grading standard of imprinting deletion and copy number abnormality, Figure 5(c) shows the distribution range and grading standard of imprinted deletion and copy number abnormality in 37 cases of gastric tumor pathological sections of the imprinted gene Z16, Figure 5(d) ) is the distribution range and grading standard of imprinting deletion and copy number abnormality in 37 cases of gastric tumor pathological sections applied to imprinted gene Z3. The distribution range and grading standard of copy number abnormality, Figure 5(f) is the distribution range and grading standard of imprinted deletion and copy number abnormality in 37 cases of gastric tumor pathological sections of imprinted gene Z5, Figure 5(g) is the imprinted gene The distribution range and grading standard of imprinting deletion and copy number abnormality in the pathological sections of 37 cases of gastric tumors using Z6 Distribution range and grading standard, Figure 5(i) is the distribution range and grading standard of imprinted deletion and copy number abnormality in 37 cases of gastric tumor pathological sections when imprinted gene Z10 is applied, Figure 5(j) is the application of imprinted gene Z13 in 37 cases The distribution range and grading standard of imprinting deletion and copy number abnormality in the pathological section of gastric tumor, where LOI is the gene expression level of imprinted gene deletion, CNV is the gene expression level of imprinted gene copy number abnormality, and TE is the total expression level of imprinted gene ;

Figure 6(a) is the expression status of 10 genes in the pathological section of grade 0 esophageal tumor, Figure 6(b) is the expression status of 10 genes in the pathological section of grade I esophageal cancer, and Figure 6(c) is grade II The expression status of 10 genes in the pathological section of esophageal cancer, Figure 6(d) shows the expression status of 10 genes in the pathological section of grade III esophageal cancer, and Figure 6(e) shows the expression status of 10 genes in the pathological section of grade IV esophageal cancer gene expression status;

Figure 7(a) is the intensity of imprinting deletion of imprinted genes Z1, Z11 and Z16 in esophageal cancer, Figure 7(b) is the intensity of imprinted genes Z1, Z11 and Z16 in esophageal cancer copy number abnormality, Figure 7(c) is the intensity of the total expression of the imprinted genes Z1, Z11 and Z16 in esophageal cancer, Figure 7(d) is the intensity of the imprinted deletion of the imprinted genes Z3, Z4, Z5, Z6, Z8, Z10 and Z13 in esophageal cancer, Figure 7 (e) is the intensity of the copy number abnormalities of imprinted genes Z3, Z4, Z5, Z6, Z8, Z10 and Z13 in esophageal cancer, Figure 7(f) is the imprinted genes Z3, Z4, Z5, Z6, Z8, Z10 and Z13 The intensity of the total expression of esophageal cancer, where LOI is the expression of missing imprinted genes, CNV is the expression of imprinted gene copy number abnormalities, and TE is the total expression of imprinted genes;

Figure 8(a) is the intensity of imprinted deletion, copy number abnormality and total expression of imprinted gene Z1, Figure 8(b) is the intensity of imprinted deletion, copy number abnormality and total expression of imprinted gene Z11, Figure 8(c) is the intensity of imprinted deletion, copy number abnormality and total expression of imprinted gene Z16, Figure 8(d) is the intensity of imprinted deletion, copy number abnormality and total expression of imprinted gene Z3, and Figure 8(e) is the imprinted deletion of imprinted gene Z4 , copy number abnormality and total expression intensity, Figure 8(f) is the intensity of imprinting deletion, copy number abnormality and total expression of imprinted gene Z5, Figure 8(g) is imprinted gene Z6 imprinting deletion, copy number abnormality and total expression The intensity of expression level, Figure 8(h) is the intensity of imprinting deletion, copy number abnormality and total expression level of imprinted gene Z8, Figure 8(i) is the intensity of imprinted gene Z10 imprinting deletion, copy number abnormality and total expression level. 8(j) is the intensity of imprinting deletion, copy number abnormality and total expression level of imprinted gene Z13, where LOI is the expression level of imprinted gene deletion gene, CNV is the gene expression level of imprinted gene copy number abnormality, and TE is the total expression level of imprinted gene quantity;

Figure 9(a) shows the distribution range and grading standard of imprinted deletion and copy number abnormality applied to 57 cases of esophageal tumor pathological sections of imprinted gene Z1, and Figure 9(b) shows the application of imprinted gene Z11 to 57 cases of esophageal tumor pathological sections , the distribution range and grading standard of imprinted deletion and copy number abnormality, Figure 9(c) shows the distribution range and grading standard of imprinted deletion and copy number abnormality in 57 cases of esophageal tumor pathological sections of the imprinted gene Z16, Figure 9(d) ) is the distribution range and grading standard of imprinting deletion and copy number abnormality in 57 cases of esophageal tumor pathological sections applied to imprinted gene Z3. Figure 9(e) is the application of imprinted gene Z4 to 57 cases of esophageal tumor pathological sections. The distribution range and grading standard of copy number abnormality, Figure 9(f) is the distribution range and grading standard of imprinted deletion and copy number abnormality in 57 cases of esophageal tumor pathological sections of imprinted gene Z5, Figure 9(g) is the imprinted gene The distribution range and grading standard of imprinting deletion and copy number abnormality in 57 cases of esophageal tumor pathological sections using Z6 Distribution range and grading standard, Figure 9(i) is the distribution range and grading standard of imprinted deletion and copy number abnormality in 57 cases of esophageal tumor pathological sections applied to imprinted gene Z10, Figure 9(j) is the application of imprinted gene Z13 to 57 The distribution range and grading standard of imprinting deletion and copy number abnormality in the pathological section of esophageal tumor, where LOI is the expression level of imprinted gene deletion genes, CNV is the gene expression level of imprinted gene copy number abnormality, and TE is the total expression level of imprinted genes .

Detailed ways

In order to further illustrate the technical means adopted by the present invention and its effects, the technical solutions of the present invention are further described below with reference to the accompanying drawings and specific embodiments, but the present invention is not limited to the scope of the embodiments.

Example 1 Imprinted gene analysis of gastric tumors

The method for detecting an imprinted gene comprises the following steps:

(1) Obtain tissue and cell sections (10 microns) of gastric tumors, put them in 10% neutral formalin solution for fixation to prevent RNA degradation, fixation time is 24 hours, paraffin-embedded (FFPE), the described The slides need to be removed with a positive charge, and the slides should be baked in a 40°C oven for more than 3 hours;

(2) Perform dewaxing treatment according to the sample processing method of RNASCope, block endogenous peroxidase activity in the sample, enhance permeability and expose RNA molecules;

(3) Design probes: design specific primers according to the imprinted gene sequence;

The designed probes are based on the imprinted genes Z1 (Gnas), Z3 (Peg10), Z4 (Igf2r), Z5 (Mest), Z6 (Plagl1), Z8 (Dcn), Z10 (Gatm), Z11 (Grb10), Z13 (Sgce) and Z16 (Snrpn/Snurf), specifically, a sequence was selected in the endoroton of each gene as a probe, and the specific probe was designed by Advanced CellDiagnostics.

(4) performing RNA SCope in situ hybridization with the probe of step (3) and the sample to be tested by the kit;

(5) Signal amplification and hematoxylin staining to analyze the expression of imprinted genes with microscope imaging;

The formulas for calculating the total expression of imprinted genes, the missing expression of imprinted genes and the abnormal expression of imprinted gene copy number in the model are as follows:

Total expression = (b+c+d)/(a+b+c+d)×100%;

Total expression of normal imprinted genes=b/(b+c+d)×100%;

Imprinted gene deletion gene expression (LOI)=c/(b+c+d)×100%;

Gene expression with abnormal copy number of imprinted genes (CNV)=d/(b+c+d)×100%;

Among them, a, b, c, and d are shown in Figure 1. The a is the nucleus of the cell where there is no marker in the nucleus after hematoxylin staining, and the imprinted gene is not expressed; the b is the cell after hematoxylin staining. There is one red/brown mark in the nucleus, the nucleus with the imprinted gene; the c is the nucleus with two red/brown marks in the nucleus after the cells are stained with hematoxylin, and the imprinted gene is missing; the d is the cell with hematoxylin. After staining, there are more than two red/brown markers in the nucleus, imprinting nuclei with abnormal copy number of genes.

As can be seen from Fig. 2(a)-Fig. 2(e), in samples from grade 0 to grade IV, imprinting loss (two signal points in the nucleus) and copy number abnormality (three or more signal points in the nucleus) ) increased gradually with the degree of malignancy.

Example 2 Imprinted gene analysis of gastroscopic biopsy samples

The gastroscopic biopsy sample is obtained by taking out suspicious lesion tissue under gastroscope, and fixing with 10% neutral formalin solution for more than 24 hours. Other detection methods are the same as those in Example 1.

It can be seen from Figure 3(a)-Figure 3(f) that the response sensitivity of each gene of Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13, Z16 to gastric cancer or corresponding to gastric cancer expression The intensity and status of imprint deletions vary.

The specific sensitivity of each imprinted gene to gastric cancer is shown in Fig. 4(a)-Fig. 4(j). From Fig. 4(a)-Fig. 4(c), it can be seen that the imprinting deletion and expression increase of the imprinted gene Z1 are in the The malignant potential stage begins to appear, and rapidly rises to a very high level during the development of early to advanced gastric cancer. The copy number abnormality of the imprinted gene Z1 begins to appear in the malignant potential stage, rapidly increases in early and intermediate gastric cancer, and rises in the advanced gastric cancer stage. The speed slowed down; the imprinted deletion of the imprinted gene Z11 began to appear in the malignant potential stage, rapidly increased in the early gastric cancer stage, slowed down in the mid-stage gastric cancer, and continued to rise to a higher level in the advanced gastric cancer stage, and the copy number of the imprinted gene Z11 was abnormal. The imprinted gene Z16, with its imprinted deletion, copy number abnormality and increased expression, increased rapidly in the malignant potential and early gastric cancer stages. , but there is a significant decline in the stage of middle and advanced gastric cancer;

From Figure 4(d)-Figure 4(j), it can be seen that the imprinting deletion, copy number abnormality and increased expression of the imprinted gene Z3 begin to appear in the early gastric cancer stage, continue to rise in the mid-stage gastric cancer, and increase rapidly in the advanced gastric cancer Slow down; the imprinted deletion of imprinted gene Z4 begins to appear in the early gastric cancer stage, rises rapidly in the intermediate gastric cancer stage, and slows down in the advanced gastric cancer stage. The rate of increase slowed down, and the increase in the expression of imprinted gene Z4 began to appear in the malignant potential stage, rapidly increased in the early gastric cancer stage, did not increase significantly in the intermediate gastric cancer stage, and continued to increase in the advanced gastric cancer stage; the imprinting of the imprinted gene Z5 was lost in the early gastric cancer stage. Rapidly rising, the rate of increase slowed down in the mid-stage gastric cancer, and continued to rise to a higher level in the advanced gastric cancer stage. The copy number abnormality of the imprinted gene Z5 began to appear in the early gastric cancer stage, rapidly increased in the mid-stage gastric cancer, and increased rapidly in the advanced gastric cancer stage. Slowly, the increase in the expression of imprinted gene Z5 began to appear in early gastric cancer, and gradually increased to a higher level in intermediate and advanced gastric cancer; the imprinted deletion of imprinted gene Z6 began to appear in early gastric cancer, and continued to rise in intermediate gastric cancer. The rate of increase in advanced gastric cancer slowed down, and the abnormal copy number of imprinted gene Z6 began to appear in early gastric cancer, slowed down in mid-stage gastric cancer, and continued to increase in advanced gastric cancer, and the expression of imprinted gene Z6 increased in early gastric cancer. , increased slowly in mid-stage and advanced gastric cancer; the imprinting deletion and increased expression of the imprinted gene Z8 began to appear in early gastric cancer, slowly increased in mid-stage gastric cancer, and increased at an advanced stage of gastric cancer, but the level was still not high, the imprinted gene The copy number abnormality of Z8 increased rapidly in the early gastric cancer stage, but did not continue to increase in the intermediate and advanced gastric cancer; the imprinted deletion, copy number abnormality and increased expression of the imprinted gene Z10 began to appear in the malignant potential stage, and in early to advanced gastric cancer It gradually increased to a higher level during the development process; the imprinted deletion and increased expression of the imprinted gene Z13 began to appear in the malignant potential stage, gradually increased in early and mid-stage gastric cancer, and slowed down in advanced gastric cancer. The copy number of the imprinted gene Z13 Abnormalities began to appear in the malignant potential stage, and the rate of increase slowed down in early gastric cancer, increased in mid-stage gastric cancer, and slowed again in advanced gastric cancer.

Example 3 Imprinted gene analysis of 37 gastric tumor samples

Tissues including gastroscopic biopsy samples (10 microns) were obtained from 37 gastric tumor patients, and the detection method was the same as that of Example 1.

From Figure 5(a)-Figure 5(j), it can be seen that the imprinting deletions of ten probes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 in 37 gastric tumor tissue samples The proportion of abnormal copy number shows a distribution from low to high. According to the distribution trend of different probes, we calculated the grading standard shown by the dotted line in the figure, and can classify the imprint deletion and copy number abnormality of each probe from low to low. To high is divided into 5 levels.

The specific classification is as follows:

As can be seen from Figure 5(a), for the imprinted gene Z1, any one of the missing expression of imprinted genes is less than 16%, the abnormal expression of imprinted gene copy number is less than 2.5%, or the total expression of imprinted genes is less than 35% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 2.5-4%, or the total expression of imprinted genes is 35-40% or The combination of at least two is grade I, any one of or at least 20-25% missing imprinted gene expression, 4-6.5% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, any one or at least two of the missing expression of imprinted genes is 25-30%, the abnormal expression of imprinted gene copy number is 6.5-8%, or the total expression of imprinted genes is 50-65%. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 8%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 5(b), for the imprinted gene Z11, any one of the missing expression of imprinted genes is less than 16%, the abnormal expression of imprinted gene copy number is less than 1.5%, or the total expression of imprinted genes is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 5(c), for the imprinted gene Z16, any one of the missing expression of imprinted genes is less than 15%, the abnormal expression of imprinted gene copy number is less than 1.5%, or the total expression of imprinted genes is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 15-20%, the abnormal expression of imprinted gene copy number is 1.5-4%, or the total expression of imprinted genes is 30-40% or A combination of at least two of them is grade I, any one of or at least 20-25% of imprinted gene missing expression, 4-7% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, any one or at least two of the missing expression of imprinted genes is 25-30%, the abnormal expression of imprinted gene copy number is 7-10%, or the total expression of imprinted genes is 50-60%. The combination of species is grade III, and the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 10%, or the total expression of imprinted genes is greater than 60%. Any one or a combination of at least two of them is grade IV;

As can be seen from Figure 5(d), for the imprinted gene Z3, any one of the missing expression of imprinted genes is less than 10%, the abnormal expression of imprinted gene copy number is less than 1%, or the total expression of imprinted genes is less than 25% Or the combination of at least two is grade 0, any one of the missing expression of imprinted genes is 10-15%, the abnormal expression of imprinted gene copy number is 1-2%, or the total expression of imprinted genes is 25-30% or The combination of at least two is grade I, any one of or at least 15-25% of imprinted gene missing expression, 2-3% abnormal imprinted gene copy number expression, or 30-45% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 3-5% abnormal imprinted gene copy number expression, or 45-55% total imprinted gene expression or at least two of them. The combination of species is grade III, and the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 5%, or the total expression of imprinted genes is greater than 55%. Any one or a combination of at least two of them is grade IV;

As can be seen from Figure 5(e), for the imprinted gene Z4, any one of the missing expression of imprinted genes is less than 16%, the abnormal expression of imprinted gene copy number is less than 1.5% or the total expression of imprinted genes is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 5(f), for the imprinted gene Z5, any one of the missing expression of imprinted genes is less than 16%, the abnormal expression of imprinted gene copy number is less than 1.5% or the total expression of imprinted genes is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 5(g), for the imprinted gene Z6, any one of the missing expression level of the imprinted gene is less than 16%, the abnormal expression level of the imprinted gene copy number is less than 1.5% or the total expression level of the imprinted gene is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 5(h), for the imprinted gene Z8, any one of the missing expression of imprinted genes is less than 10%, the abnormal expression of imprinted gene copy number is less than 1% or the total expression of imprinted genes is less than 25% Or the combination of at least two is grade 0, any one of the missing expression of imprinted genes is 10-15%, the abnormal expression of imprinted gene copy number is 1-2%, or the total expression of imprinted genes is 25-30% or The combination of at least two is grade I, any one of or at least 15-25% of imprinted gene missing expression, 2-3% abnormal imprinted gene copy number expression, or 30-45% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 3-5% abnormal imprinted gene copy number expression, or 45-55% total imprinted gene expression or at least two of them. The combination of species is grade III, and the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 5%, or the total expression of imprinted genes is greater than 55%. Any one or a combination of at least two of them is grade IV;

As can be seen from Figure 5(i), for the imprinted gene Z10, any one of the missing expression of imprinted genes is less than 16%, the abnormal expression of imprinted gene copy number is less than 1.5% or the total expression of imprinted genes is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 5(j), for the imprinted gene Z13, any one of the missing expression level of the imprinted gene is less than 10%, the abnormal expression level of the imprinted gene copy number is less than 1% or the total expression level of the imprinted gene is less than 25% Or the combination of at least two is grade 0, any one of the missing expression of imprinted genes is 10-15%, the abnormal expression of imprinted gene copy number is 1-2%, or the total expression of imprinted genes is 25-30% or The combination of at least two is grade I, any one of or at least 15-25% of imprinted gene missing expression, 2-3% abnormal imprinted gene copy number expression, or 30-45% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 3-5% abnormal imprinted gene copy number expression, or 45-55% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 5%, or the total expression of imprinted genes is greater than 55%, or the combination of at least two is grade IV.

From the comprehensive analysis of these 37 gastric tumor samples, it can be concluded that:

To judge the benign and malignant degree of gastric tumor, it can be divided into benign tumor, gastric cancer potential, early gastric cancer, intermediate gastric cancer and advanced gastric cancer:

The result of judging the benign and malignant degree of gastric tumor is that the imprinted gene deletion expression amount and imprinted gene copy number abnormal expression amount of imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 are all less than 1 Level I, imprinted gene deletion expression level of no more than 1 imprinted gene among imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16, and imprinted genes Z1, Z3, Z4, Z5 , Z6, Z8, Z10, Z11, Z13 and Z16, the abnormal expression of imprinted gene copy number of no more than 1 imprinted gene is any one of grade I, then it is a benign tumor;

The result of judging the benign and malignant degree of gastric tumor is that the expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level I, and the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 had at least 2 imprinted genes with abnormal expression of imprinted gene copy number at grade I, and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10 , Z11 and Z13 imprinted gene deletion expression level of no more than 1 imprinted gene level I and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 no more than 1 imprinted gene The abnormal expression level of imprinted gene copy number is level I and the imprinted gene Z16 has missing expression level of imprinted gene or the abnormal expression level of imprinted gene copy number is level I or above, or imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10 , Z11 and Z13 with no more than 1 imprinted gene deletion expression level II and imprinted with no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 If the abnormal expression of gene copy number is any one of grade II, it is gastric cancer potential;

The result of judging the benign and malignant degree of gastric tumor is that the expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level II, and the imprinted gene expression level is II. At least 2 imprinted genes in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 have abnormal expression levels of imprinted gene copy number at level II or imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Imprinted gene deletion expression level of no more than 1 imprinted gene in Z10, Z11 and Z13 is grade III and no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13. If the abnormal expression level of imprinted gene copy number is any one of grade III, it is early gastric cancer;

The result of judging the benign and malignant degree of gastric tumor is that the expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is level III, and the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 have at least 2 imprinted genes with abnormal expression of imprinted gene copy number at level III or imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10 , Z11 and Z13 imprinted gene deletion expression level of no more than 1 imprinted gene level IV and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 imprinted no more than 1 imprinted gene If the abnormal expression of gene copy number is any of the IV grades, it is mid-stage gastric cancer;

The result of judging the benign and malignant degree of gastric tumor is that the imprinted gene deletion expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is grade IV or the imprinted gene Z1 , Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 at least 2 imprinted genes in the abnormal expression of imprinted gene copy number grade IV, it is advanced gastric cancer.

Example 4 Imprinted gene analysis of esophageal tumors

The tissue and cell sections (10 microns) of the esophageal tumor were obtained, and the detection method was the same as that in Example 1.

As can be seen from Fig. 6(a)-Fig. 6(e), in samples from grade 0 to grade IV, imprinting loss (two signal points in the nucleus) and copy number abnormality (three or more signal points in the nucleus) ) increased gradually with the degree of malignancy.

Example 5 Imprinted gene analysis of gastroscopic biopsy samples of esophagus

The gastroscopic biopsy sample of the esophagus is obtained by taking out the suspicious diseased tissue of the esophagus under a gastroscope, and fixing it with 10% neutral formalin solution for more than 24 hours. Other detection methods are the same as those in Example 1.

From Figure 7(a)-Figure 7(f), it can be seen that the sensitivity of each gene of Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13, Z16 to esophageal cancer or corresponding to esophageal cancer The strength and status of imprinted deletions expressed by cancers varied.

The specific sensitivity of each imprinted gene to esophageal cancer is shown in Figure 8(a)-Figure 8(j). It can be seen from Figure 8(a)-Figure 8(c) that the imprinting deletion of the imprinted gene Z1 is in the malignant potential stage. It began to appear, did not increase significantly in early esophageal cancer, and gradually increased to a very high level in intermediate and advanced esophageal cancer. The abnormal copy number of imprinted gene Z1 gradually increased from malignant potential to advanced esophageal cancer, and the expression level of imprinted gene Z1 The increase rises slowly during the development of esophageal cancer; the imprinted deletion of the imprinted gene Z11 begins to appear in the malignant potential stage, does not increase significantly in the early and middle esophageal cancer stages, and increases significantly in the advanced esophageal cancer stage, the copy of the imprinted gene Z11 The number of abnormalities increased rapidly in the malignant potential stage, and gradually slowed down during the development of early to advanced esophageal cancer. The increase in the expression of imprinted gene Z11 did not increase significantly in the malignant potential to mid-stage esophageal cancer stages, but in advanced esophageal cancer. There was a significant increase; the imprinted deletion, copy number abnormality and increased expression of the imprinted gene Z16 began to appear in the malignant potential stage, rapidly increased in the early and middle stage of esophageal cancer, and slowed down in the advanced stage of esophageal cancer;

It can be seen from Figure 8(d)-Figure 8(j) that the imprinted deletion, copy number abnormality and increased expression of the imprinted gene Z3 started to increase in the intermediate stage of esophageal cancer, and continued to increase in advanced esophageal cancer, but the levels were still not high High; the imprinted deletion of imprinted gene Z4 begins to appear in the intermediate stage of esophageal cancer, but the rate of increase slows down in the advanced stage of esophageal cancer, and the copy number abnormalities and increased expression of imprinted genes begin to appear in the intermediate stage of esophageal cancer and continue in advanced esophageal cancer Rising; imprinted deletion, copy number abnormality and increased expression of imprinted gene Z5 began to increase in the intermediate stage of esophageal cancer and continued to increase in advanced esophageal cancer, but the level was still not high; the imprinted deletion of imprinted gene Z6 started in the intermediate stage of esophageal cancer Appeared, accelerated in advanced esophageal cancer, but the level was still not high. The abnormal copy number and increased expression of imprinted gene Z6 began to appear in the malignant potential stage, and slowly increased with the development of esophageal cancer, and the level remained at the advanced esophageal cancer stage. Not high; imprinted deletion and copy number abnormalities of imprinted gene Z8 begin to appear at the malignant potential stage, rapidly increase in early and mid-stage esophageal cancer, and slow down at the advanced esophageal cancer stage, and the expression of imprinted gene Z8 increases in the malignant potential stage began to appear, but did not increase significantly during the development of esophageal cancer; the imprinting deletion of imprinted gene Z10 began to appear in the intermediate esophageal cancer stage, and did not continue to increase in the advanced esophageal cancer stage, and the copy number abnormality of the imprinted gene Z10 began in the intermediate esophageal cancer stage appeared, and continued to rise in advanced esophageal cancer, but the level was not high. The increase in the expression of imprinted gene Z10 began to appear in the malignant potential stage, but did not increase significantly during the development of esophageal cancer; the imprinted deletion of imprinted gene Z13 was found in early esophageal cancer. It began to appear in the middle and advanced stages of esophageal cancer, but the level was not high. The abnormal copy number and increased expression of the imprinted gene Z13 increased rapidly in the intermediate stage of esophageal cancer and slowed down in the advanced stage of esophageal cancer.

Example 6 Imprinted gene analysis of 57 esophageal tumor samples

The tissues of 57 patients with esophageal tumor including esophagus gastroscopic biopsy samples (10 microns) were obtained, and the detection method was the same as that of Example 1.

As can be seen from Fig. 9(a)-Fig. 9(j), the imprinting deletions of ten probes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 in 57 esophageal tumor tissue samples The proportion of abnormal copy number shows a distribution from low to high. According to the distribution trend of different probes, we calculated the grading standard shown by the dotted line in the figure, and can classify the imprint deletion and copy number abnormality of each probe from low to low. To high is divided into 5 levels.

The specific classification is as follows:

As can be seen from Figure 9(a), for the imprinted gene Z1, any one of the missing expression of imprinted genes is less than 16%, the abnormal expression of imprinted gene copy number is less than 2.5%, or the total expression of imprinted genes is less than 35% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 2.5-4%, or the total expression of imprinted genes is 35-40% or The combination of at least two is grade I, any one of or at least 20-25% missing imprinted gene expression, 4-6.5% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, any one or at least two of the missing expression of imprinted genes is 25-30%, the abnormal expression of imprinted gene copy number is 6.5-8%, or the total expression of imprinted genes is 50-65%. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 8%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 9(b), for the imprinted gene Z11, any one of the missing expression of imprinted genes is less than 16%, the abnormal expression of imprinted gene copy number is less than 1.5%, or the total expression of imprinted genes is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 9(c), for the imprinted gene Z16, any one of the missing expression level of the imprinted gene is less than 10%, the abnormal expression level of the imprinted gene copy number is less than 1% or the total expression level of the imprinted gene is less than 25% Or the combination of at least two is grade 0, any one of the missing expression of imprinted genes is 10-15%, the abnormal expression of imprinted gene copy number is 1-2%, or the total expression of imprinted genes is 25-30% or The combination of at least two of them is grade I, and the missing expression of imprinted genes is 15-25%, the abnormal expression of imprinted gene copy number is 2-4%, or the total expression of imprinted genes is 30-40%. Any one or at least The combination of the two is grade II, any one or at least two of the missing expression of imprinted genes is 25-30%, the abnormal expression of imprinted gene copy number is 4-6%, or the total expression of imprinted genes is 40-55%. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 55%, or the combination of at least two is grade IV;

As can be seen from Figure 9(d), for the imprinted gene Z3, any one of the missing expression of imprinted genes is less than 10%, the abnormal expression of imprinted gene copy number is less than 1% or the total expression of imprinted genes is less than 25% Or the combination of at least two is grade 0, any one of the missing expression of imprinted genes is 10-15%, the abnormal expression of imprinted gene copy number is 1-2%, or the total expression of imprinted genes is 25-30% or The combination of at least two is grade I, any one of or at least 15-25% of imprinted gene missing expression, 2-3% abnormal imprinted gene copy number expression, or 30-45% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 3-5% abnormal imprinted gene copy number expression, or 45-55% total imprinted gene expression or at least two of them. The combination of species is grade III, and the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 5%, or the total expression of imprinted genes is greater than 55%. Any one or a combination of at least two of them is grade IV;

As can be seen from Figure 9(e), for the imprinted gene Z4, any one of the missing expression level of the imprinted gene is less than 16%, the abnormal expression level of the imprinted gene copy number is less than 1.5%, or the total expression level of the imprinted gene is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 9(f), for the imprinted gene Z5, any one of the missing expression of imprinted genes is less than 16%, the abnormal expression of imprinted gene copy number is less than 1.5%, or the total expression of imprinted genes is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 9(g), for the imprinted gene Z6, any one of the missing expression of the imprinted gene is less than 16%, the abnormal expression of the imprinted gene copy number is less than 1.5%, or the total expression of the imprinted gene is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 9(h), for the imprinted gene Z8, any one of the missing expression level of the imprinted gene is less than 16%, the abnormal expression level of the imprinted gene copy number is less than 2.5% or the total expression level of the imprinted gene is less than 35% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 2.5-4%, or the total expression of imprinted genes is 35-40% or The combination of at least two is grade I, any one of or at least 20-25% missing imprinted gene expression, 4-6.5% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, any one or at least two of the missing expression of imprinted genes is 25-30%, the abnormal expression of imprinted gene copy number is 6.5-8%, or the total expression of imprinted genes is 50-65%. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 8%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 9(i), for the imprinted gene Z10, any one of the missing expression level of the imprinted gene is less than 16%, the abnormal expression level of the imprinted gene copy number is less than 1.5%, or the total expression level of the imprinted gene is less than 30% Or a combination of at least two of them is grade 0, any one of the missing expression of imprinted genes is 16-20%, the abnormal expression of imprinted gene copy number is 1.5-2.5%, or the total expression of imprinted genes is 30-40% or The combination of at least two is grade I, any one or at least 20-25% of imprinted gene missing expression, 2.5-4% abnormal imprinted gene copy number expression, or 40-50% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 4-6% abnormal imprinted gene copy number expression, or 50-65% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 6%, or the total expression of imprinted genes is greater than 65%, or the combination of at least two is grade IV;

As can be seen from Figure 9(j), for the imprinted gene Z13, any one of the missing expression level of the imprinted gene is less than 10%, the abnormal expression level of the imprinted gene copy number is less than 1%, or the total expression level of the imprinted gene is less than 25% Or the combination of at least two is grade 0, any one of the missing expression of imprinted genes is 10-15%, the abnormal expression of imprinted gene copy number is 1-2%, or the total expression of imprinted genes is 25-30% or The combination of at least two is grade I, any one of or at least 15-25% of imprinted gene missing expression, 2-3% abnormal imprinted gene copy number expression, or 30-45% total imprinted gene expression The combination of the two is grade II, with either 25-30% missing imprinted gene expression, 3-5% abnormal imprinted gene copy number expression, or 45-55% total imprinted gene expression or at least two of them. The combination of species is grade III, and any one of the missing expression of imprinted genes is greater than 30%, the abnormal expression of imprinted gene copy number is greater than 5%, or the total expression of imprinted genes is greater than 55%, or the combination of at least two is grade IV.

From the comprehensive analysis of these 57 esophageal tumor samples, it can be concluded that:

Judging the benign and malignant degree of esophageal tumor is divided into benign tumor, esophageal cancer potential, early esophageal cancer, intermediate esophageal cancer and advanced esophageal cancer:

The result of judging the benign and malignant degree of esophageal tumor is that the imprinted gene deletion expression amount and imprinted gene copy number abnormal expression amount of imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 are all less than 1 Level I, imprinted gene deletion expression level of no more than 1 imprinted gene among imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16, and imprinted genes Z1, Z3, Z4, Z5 , Z6, Z8, Z10, Z11, Z13 and Z16, the abnormal expression of imprinted gene copy number of no more than 1 imprinted gene is any one of grade I, then it is a benign tumor;

The result of judging the benign and malignant degree of esophageal tumor is that the imprinted gene deletion expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is level I, Imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 abnormal expression level I or imprinted gene Z1, Z3, Z4, Z5, Z6 , Z8, Z10, Z11, Z13 and Z16 imprinted gene deletion expression level of no more than 1 imprinted gene level II and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 If the abnormal expression of imprinted gene copy number of no more than 1 imprinted gene is any one of grade II, it is esophageal cancer potential;

The result of judging the benign and malignant degree of esophageal tumor is that the expression level of at least two imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is level II, Imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 at least 2 imprinted genes with abnormal expression of imprinted gene copy number at level II or imprinted genes Z1, Z3, Z4, Z5, Z6 , Z8, Z10, Z11, Z13, and Z16 with no more than 1 imprinted gene deletion expression level III and among the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 If the abnormal expression of imprinted gene copy number of no more than one imprinted gene is any one of grade III, it is early esophageal cancer;

The result of judging the benign and malignant degree of esophageal tumor is that the expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is level III, Imprinted gene copy number of at least 2 imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 abnormal expression level III or imprinted gene Z1, Z3, Z4, Z5, Z6 , Z8, Z10, Z11, Z13 and Z16 imprinted gene deletion expression level of no more than 1 imprinted gene level IV and imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 If the abnormal expression of imprinted gene copy number of no more than one imprinted gene is any one of grade IV, it is intermediate-stage esophageal cancer;

The result of judging the benign and malignant degree of esophageal tumor is that the imprinted gene deletion expression level of at least 2 imprinted genes in the imprinted genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade IV or imprinted The abnormal expression of imprinted gene copy number of at least 2 imprinted genes in the genes Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is grade IV, which is advanced esophageal cancer.

To sum up, the detection model and system of the present invention express the performance of imprinting deletion in the samples of patients with esophagus and gastric tumors in an intuitive way. The changes of imprinted (trace) genes can be accurately detected, and a quantitative model can be provided, making a great contribution to the diagnosis of esophageal and gastric tumors.

The applicant declares that the present invention illustrates the detailed method of the present invention through the above-mentioned embodiments, but the present invention is not limited to the above-mentioned detailed method, that is, it does not mean that the present invention must rely on the above-mentioned detailed method to be implemented. Those skilled in the art should understand that any improvement of the present invention, the equivalent replacement of each raw material of the product of the present invention, the addition of auxiliary components, the selection of specific methods, etc., all fall within the protection scope and disclosure scope of the present invention.

Claims (10)

1. a kind of for esophageal tumor and/or the imprinted gene hierarchy model of stomach neoplasm, which is characterized in that the model passes through meter Total expression quantity of imprinted gene loss of expression amount, imprinted gene copy number unconventionality expression amount and imprinted gene is calculated in esophageal tumor And/or the variation in stomach neoplasm is classified the expression status of imprinted gene；

Wherein, the imprinted gene be Z1, Z11 or Z16 in any one or at least two combination, the imprinted gene Z1 It is Grb10 for Gnas, the imprinted gene Z11, the imprinted gene Z16 is Snrpn/Snurf.

2. imprinted gene hierarchy model according to claim 1, which is characterized in that the model calculates the side of imprinted gene Method is as follows:

Any one in Z1, Z11 or Z16 is calculated, any one in preferably Z1 or Z11；

When preferably, for detecting esophageal tumor, the imprinted gene that the model calculates is Z1；

When preferably, for detecting stomach neoplasm, the imprinted gene that the model calculates is Z11；

Preferably, the method that the model calculates imprinted gene are as follows: calculate any two in Z1, Z11 or Z16.

3. imprinted gene hierarchy model according to claim 1 or 2, which is characterized in that the imprinted gene further include Z3, In Z4, Z5, Z6, Z8, Z10 or Z13 any one or at least two combination；Wherein, the imprinted gene Z3 is Peg10, The imprinted gene Z4 is Igf2r, and the imprinted gene Z5 is Mest, and the imprinted gene Z6 is Plagl1, the marking base Because Z8 is Dcn, the imprinted gene Z10 is Gatm, and the imprinted gene Z13 is Sgce；

Preferably, the method that the model calculates imprinted gene are as follows: calculate the combination of imprinted gene, calculate Z1, Z3, Z4, Z5, The combination of ten imprinted genes of Z6, Z8, Z10, Z11, Z13 and Z16.

4. imprinted gene hierarchy model according to any one of claim 1-3, which is characterized in that the calculating marking base The formula of total expression quantity of cause, imprinted gene loss of expression amount and imprinted gene copy number unconventionality expression amount is as follows:

Total expression quantity=(b+c+d)/(a+b+c+d) × 100%；

Total expression quantity=the b/ (b+c+d) × 100% of normal imprinted gene；

Imprinted gene missing gene expression quantity=c/ (b+c+d) × 100%；

Gene expression amount=d/ (b+c+d) × 100% of imprinted gene copy number exception；

Wherein, a is that after cell is carried out haematoxylin dyeing, label is not present in nucleus, what imprinted gene was not expressed Nucleus；The b is after cell is carried out haematoxylin dyeing, and there are a red/brown mark in nucleus, imprinted gene is deposited Nucleus；The c is after cell is carried out haematoxylin dyeing, and there are two red/brown marks, marking bases in nucleus Because of the nucleus of missing；The d is there are more than two red/brown marks in nucleus after cell is carried out haematoxylin dyeing Note, the nucleus of imprinted gene copy number exception.

5. imprinted gene hierarchy model described in any one of -4 according to claim 1, which is characterized in that the imprinted gene lacks It loses expression quantity, imprinted gene copy number unconventionality expression amount and the total expression quantity of imprinted gene and is divided into five different grades；

Preferably, five different grades are ten prints for Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 Imprinted gene loss of expression amount, imprinted gene copy number unconventionality expression amount and the total expression quantity of imprinted gene of note gene carry out respectively The five different grades divided；

Preferably, the imprinted gene loss of expression amount, imprinted gene copy number unconventionality expression amount and total expression quantity for Z1 The five different grades divided are as follows:

0 grade: the imprinted gene loss of expression amount of the imprinted gene Z1 is copied less than the imprinted gene of 16%, the imprinted gene Z1 Shellfish number unconventionality expression amount less than 2.5% or total expression quantity of the imprinted gene Z1 less than 35% in any one or at least two The combination of kind；

I grades: imprinted gene Z1 imprinted gene loss of expression amount is the imprinted gene of 16-20%, the imprinted gene Z1 Copy number unconventionality expression amount be 2.5-4% or total expression quantity of the imprinted gene Z1 be in 35-40% any one or extremely Few two kinds of combination；

II grades: imprinted gene Z1 imprinted gene loss of expression amount is the imprinted gene of 20-25%, the imprinted gene Z1 Copy number unconventionality expression amount be 4-6.5% or total expression quantity of the imprinted gene Z1 be in 40-50% any one or extremely Few two kinds of combination；

III level: the imprinted gene loss of expression amount of the imprinted gene Z1 is the marking base of 25-30%, the imprinted gene Z1 Because copy number unconventionality expression amount be 6.5-8% or total expression quantity of the imprinted gene Z1 be in 50-65% any one or At least two combination；

IV grades: imprinted gene Z1 imprinted gene loss of expression amount is greater than the imprinted gene of 30%, the imprinted gene Z1 Total expression quantity of the copy number unconventionality expression amount greater than the 8% or imprinted gene Z1 is greater than any one or at least two in 65% The combination of kind；

Preferably, the imprinted gene loss of expression amount, imprinted gene copy number unconventionality expression amount and summary table for Z3 and Z13 The five different grades divided up to amount are as follows:

0 grade: the imprinted gene loss of expression amount of the imprinted gene Z3 and Z13 is less than 10%, the imprinted gene Z3's and Z13 Imprinted gene copy number unconventionality expression amount less than 1% or total expression quantity of the imprinted gene Z3 and Z13 less than 25% in appoint It anticipates a kind of or at least two combinations；

I grades: the imprinted gene Z3 and Z13 imprinted gene loss of expression amount is 10-15%, the imprinted gene Z3 and Z13 Imprinted gene copy number unconventionality expression amount be 1-2% or total expression quantity of the imprinted gene Z3 and Z13 is in 25-30% Any one or at least two combination；

II grades: the imprinted gene Z3 and Z13 imprinted gene loss of expression amount is 15-25%, the imprinted gene Z3 and Z13 Imprinted gene copy number unconventionality expression amount be 2-3% or total expression quantity of the imprinted gene Z3 and Z13 is in 30-45% Any one or at least two combination；

III level: the imprinted gene loss of expression amount of the imprinted gene Z3 and Z13 be 25-30%, the imprinted gene Z3 and The imprinted gene copy number unconventionality expression amount of Z13 is 3-5% or total expression quantity of the imprinted gene Z3 and Z13 is 45-55% In any one or at least two combination；

IV grades: the imprinted gene Z3 and Z13 imprinted gene loss of expression amount is greater than 30%, the imprinted gene Z3 and Z13 Imprinted gene copy number unconventionality expression amount greater than the 5% or imprinted gene Z3 and Z13 total expression quantity be greater than 55% in Any one or at least two combination；

Preferably, the imprinted gene loss of expression amount for Z4, Z5, Z6, Z10 and Z11, imprinted gene copy number exception table The five different grades divided up to amount with total expression quantity are as follows:

0 grade: the imprinted gene loss of expression amount of described imprinted gene Z4, Z5, Z6, Z10 and Z11 are less than the 16%, marking base Because Z4, Z5, Z6, Z10 and Z11 imprinted gene copy number unconventionality expression amount less than 1.5% or the imprinted gene Z4, Z5, Z6, Total expression quantity of Z10 and Z11 less than 30% in any one or at least two combination；

I grades: the imprinted gene loss of expression amount of described imprinted gene Z4, Z5, Z6, Z10 and Z11 are 16-20%, the marking base Because Z4, Z5, Z6, Z10 and Z11 imprinted gene copy number unconventionality expression amount be the 1.5-2.5% or imprinted gene Z4, Z5, Total expression quantity of Z6, Z10 and Z11 are the combination of any one or at least two in 30-40%；

II grades: the imprinted gene loss of expression amount of described imprinted gene Z4, Z5, Z6, Z10 and Z11 are 20-25%, the marking The imprinted gene copy number unconventionality expression amount of gene Z4, Z5, Z6, Z10 and Z11 be the 2.5-4% or imprinted gene Z4, Z5, Total expression quantity of Z6, Z10 and Z11 are the combination of any one or at least two in 40-50%；

III level: the imprinted gene loss of expression amount of described imprinted gene Z4, Z5, Z6, Z10 and Z11 are 25-30%, the marking The imprinted gene copy number unconventionality expression amount of gene Z4, Z5, Z6, Z10 and Z11 be the 4-6% or imprinted gene Z4, Z5, Z6, Total expression quantity of Z10 and Z11 is the combination of any one or at least two in 50-65%；

IV grades: the imprinted gene loss of expression amount of described imprinted gene Z4, Z5, Z6, Z10 and Z11 are greater than 30%, the marking base Because Z4, Z5, Z6, Z10 and Z11 imprinted gene copy number unconventionality expression amount be greater than the 6% or imprinted gene Z4, Z5, Z6, Total expression quantity of Z10 and Z11 is greater than the combination of any one or at least two in 65%；

It is described for the imprinted gene loss of expression amount of Z16, imprinted gene copy number when preferably, for detecting esophageal tumor Five different grades that unconventionality expression amount and total expression quantity divide are as follows:

0 grade: imprinted gene of the imprinted gene loss of expression amount of the imprinted gene Z16 less than 10%, the imprinted gene Z16 Copy number unconventionality expression amount less than 1% or total expression quantity of the imprinted gene Z16 less than 25% in any one or at least Two kinds of combination；

I grades: imprinted gene Z16 imprinted gene loss of expression amount is the marking base of 10-15%, the imprinted gene Z16 Because copy number unconventionality expression amount be 1-2% or total expression quantity of the imprinted gene Z16 be in 25-30% any one or extremely Few two kinds of combination；

II grades: imprinted gene Z16 imprinted gene loss of expression amount is the marking base of 15-25%, the imprinted gene Z16 Because copy number unconventionality expression amount be 2-4% or total expression quantity of the imprinted gene Z16 be in 30-40% any one or extremely Few two kinds of combination；

III level: the imprinted gene loss of expression amount of the imprinted gene Z16 is the marking of 25-30%, the imprinted gene Z16 Gene copy number unconventionality expression amount be 4-6% or total expression quantity of the imprinted gene Z16 be in 40-55% any one or At least two combination；

IV grades: imprinted gene Z16 imprinted gene loss of expression amount is greater than the marking base of 30%, the imprinted gene Z16 Because copy number unconventionality expression amount greater than the 6% or imprinted gene Z16 total expression quantity be greater than 55% in any one or extremely Few two kinds of combination；

It is described different for the imprinted gene loss of expression amount of Z16, imprinted gene copy number when preferably, for detecting stomach neoplasm Five different grades that normal expression quantity and total expression quantity divide are as follows:

0 grade: imprinted gene of the imprinted gene loss of expression amount of the imprinted gene Z16 less than 15%, the imprinted gene Z16 Copy number unconventionality expression amount less than 1.5% or total expression quantity of the imprinted gene Z16 less than 30% in any one or extremely Few two kinds of combination；

I grades: imprinted gene Z16 imprinted gene loss of expression amount is the marking base of 15-20%, the imprinted gene Z16 Because copy number unconventionality expression amount be 1.5-4% or total expression quantity of the imprinted gene Z16 be in 30-40% any one or At least two combination；

II grades: imprinted gene Z16 imprinted gene loss of expression amount is the marking base of 20-25%, the imprinted gene Z16 Because copy number unconventionality expression amount be 4-7% or total expression quantity of the imprinted gene Z16 be in 40-50% any one or extremely Few two kinds of combination；

III level: the imprinted gene loss of expression amount of the imprinted gene Z16 is the marking of 25-30%, the imprinted gene Z16 Gene copy number unconventionality expression amount is 7-10% or total expression quantity of the imprinted gene Z16 is any one in 50-60% Or at least two combination；

IV grades: imprinted gene Z16 imprinted gene loss of expression amount is greater than the marking base of 30%, the imprinted gene Z16 Because copy number unconventionality expression amount greater than the 10% or imprinted gene Z16 total expression quantity be greater than 60% in any one or extremely Few two kinds of combination；

It is described different for the imprinted gene loss of expression amount of Z8, imprinted gene copy number when preferably, for detecting esophageal tumor Five different grades that normal expression quantity and total expression quantity divide are as follows:

0 grade: the imprinted gene loss of expression amount of the imprinted gene Z8 is copied less than the imprinted gene of 16%, the imprinted gene Z8 Shellfish number unconventionality expression amount less than 2.5% or total expression quantity of the imprinted gene Z8 less than 35% in any one or at least two The combination of kind；

I grades: imprinted gene Z8 imprinted gene loss of expression amount is the imprinted gene of 16-20%, the imprinted gene Z8 Copy number unconventionality expression amount be 2.5-4% or total expression quantity of the imprinted gene Z8 be in 35-40% any one or extremely Few two kinds of combination；

II grades: imprinted gene Z8 imprinted gene loss of expression amount is the imprinted gene of 20-25%, the imprinted gene Z8 Copy number unconventionality expression amount be 4-6.5% or total expression quantity of the imprinted gene Z8 be in 40-50% any one or extremely Few two kinds of combination；

III level: the imprinted gene loss of expression amount of the imprinted gene Z8 is the marking base of 25-30%, the imprinted gene Z8 Because copy number unconventionality expression amount be 6.5-8% or total expression quantity of the imprinted gene Z8 be in 50-65% any one or At least two combination；

IV grades: imprinted gene Z8 imprinted gene loss of expression amount is greater than the imprinted gene of 30%, the imprinted gene Z8 Total expression quantity of the copy number unconventionality expression amount greater than the 8% or imprinted gene Z8 is greater than any one or at least two in 65% The combination of kind；

It is described abnormal for the imprinted gene loss of expression amount of Z8, imprinted gene copy number when preferably, for detecting stomach neoplasm Five different grades that expression quantity and total expression quantity divide are as follows:

0 grade: the imprinted gene loss of expression amount of the imprinted gene Z8 is copied less than the imprinted gene of 10%, the imprinted gene Z8 Shellfish number unconventionality expression amount less than 1% or total expression quantity of the imprinted gene Z8 less than 25% in any one or at least two Combination；

I grades: imprinted gene Z8 imprinted gene loss of expression amount is the imprinted gene of 10-15%, the imprinted gene Z8 Copy number unconventionality expression amount be 1-2% or total expression quantity of the imprinted gene Z8 be in 25-30% any one or at least Two kinds of combination；

II grades: imprinted gene Z8 imprinted gene loss of expression amount is the imprinted gene of 15-25%, the imprinted gene Z8 Copy number unconventionality expression amount be 2-3% or total expression quantity of the imprinted gene Z8 be in 30-45% any one or at least Two kinds of combination；

III level: the imprinted gene loss of expression amount of the imprinted gene Z8 is the marking base of 25-30%, the imprinted gene Z8 Because copy number unconventionality expression amount be 3-5% or total expression quantity of the imprinted gene Z8 be in 45-55% any one or extremely Few two kinds of combination；

IV grades: imprinted gene Z8 imprinted gene loss of expression amount is greater than the imprinted gene of 30%, the imprinted gene Z8 Total expression quantity of the copy number unconventionality expression amount greater than the 5% or imprinted gene Z8 is greater than any one or at least two in 55% The combination of kind.

6. a kind of for detecting the device of esophageal tumor and/or stomach neoplasm degree of benign and malignant, which is characterized in that including such as placing an order Member:

(1) sampling unit: sample to be tested is obtained；

(2) probe design cell: according to imprinted gene sequence design specific primer；

(3) probe of step (2) and sample to be tested detection unit: are subjected in situ hybridization；

(4) analytical unit: the expression of microscope imaging analysis imprinted gene；

Wherein, the analytical unit is led to by calculating imprinted gene loss of expression amount and imprinted gene copy number unconventionality expression amount Imprinted gene hierarchy model of any of claims 1-5 is crossed, to pass through imprinted gene loss of expression amount and the marking The grade of gene copy number unconventionality expression amount judges the degree of benign and malignant of esophageal tumor and/or stomach neoplasm.

7. system according to claim 6, which is characterized in that tissue of the sample to be tested described in step (1) from people And/or cell；

Preferably, the sample to be tested is paraffin section, esophageal mucosa cast-off cells or the gastroscopic biopsy sample of tissue, preferably Esophageal mucosa cast-off cells and/or gastroscopic biopsy sample；

Preferably, the in situ hybridization uses RNAscope in-situ hybridization method；

Preferably, the RNAscope in-situ hybridization method using single channel or multi-channel color reagents box or single channel or The Fluorescence kit of the Fluorescence kit of multichannel, preferably single channel red/brown color reagents box or multichannel.

8. device according to claim 6 or 7, which is characterized in that the degree of benign and malignant of the judgement esophageal tumor is divided into Benign tumour, cancer of the esophagus potential, early esophageal cancer, mid-term cancer of the esophagus and advanced esophageal cancer；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount and imprinted gene copy number unconventionality expression amount of Z11, Z13 and Z16 are respectively less than I grades, imprinted gene The imprinted gene loss of expression amount no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is The marking base no more than 1 imprinted gene in I grades and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 It is then benign tumour because copy number unconventionality expression amount is any one situation in I grades；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11, Z13 and Z16 be I grades, imprinted gene Z1, Z3, Z4, Z5, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z6, Z8, Z10, Z11, Z13 and Z16 is I grades or the marking It is no more than the imprinted gene loss of expression amount of 1 imprinted gene in gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 For the marking no more than 1 imprinted gene in II grades and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 Gene copy number unconventionality expression amount is any one situation in II grades, then is cancer of the esophagus potential；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11, Z13 and Z16 be II grades, imprinted gene Z1, Z3, Z4, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z5, Z6, Z8, Z10, Z11, Z13 and Z16 be II grades or The imprinted gene for being no more than 1 imprinted gene in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 lacks table It is to be no more than 1 imprinted gene in III level and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 up to amount Imprinted gene copy number unconventionality expression amount be III level in any one situation, then be early esophageal cancer；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11, Z13 and Z16 be III level, imprinted gene Z1, Z3, Z4, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z5, Z6, Z8, Z10, Z11, Z13 and Z16 be III level or The imprinted gene for being no more than 1 imprinted gene in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 lacks table Up to amount for IV grade and in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 no more than 1 imprinted gene Imprinted gene copy number unconventionality expression amount is any one situation in IV grades, then is mid-term cancer of the esophagus；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, In Z11, Z13 and Z16 the imprinted gene loss of expression amount of at least two imprinted gene be IV grades or imprinted gene Z1, Z3, Z4, Z5, In Z6, Z8, Z10, Z11, Z13 and Z16 the imprinted gene copy number unconventionality expression amount of at least two imprinted gene be IV grades, then for Advanced esophageal cancer；

Preferably, the degree of benign and malignant of the judgement stomach neoplasm is divided into benign stomach neoplasm, gastric cancer potential, early carcinoma of stomach, mid-term stomach Cancer and late gastric cancer；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount and imprinted gene copy number unconventionality expression amount of Z11, Z13 and Z16 are respectively less than I grades, imprinted gene The imprinted gene loss of expression amount no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is The marking base no more than 1 imprinted gene in I grades and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 It is then benign tumour because copy number unconventionality expression amount is any one situation in I grades；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11 and Z13 be I grades, imprinted gene Z1, Z3, Z4, Z5, Z6, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z8, Z10, Z11 and Z13 is I grades, imprinted gene Z1, The imprinted gene loss of expression amount no more than 1 imprinted gene in Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is I grades and prints Remember that the imprinted gene copy number for being no more than 1 imprinted gene in gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is abnormal Expression quantity be I grades and the imprinted gene loss of expression amount of imprinted gene Z16 or imprinted gene copy number unconventionality expression amount be I grades or More than or imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 in be no more than 1 imprinted gene imprinted gene lack Losing expression quantity is no more than 1 imprinted gene in II grade and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 Imprinted gene copy number unconventionality expression amount is any one situation in II grades, then is gastric cancer potential；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11 and Z13 be II grades, imprinted gene Z1, Z3, Z4, Z5, Z6, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene in Z8, Z10, Z11 and Z13 is II grades or imprinted gene The imprinted gene loss of expression amount for being no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is III level And the imprinted gene copy number no more than 1 imprinted gene in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 Unconventionality expression amount is any one situation in III level, then is early carcinoma of stomach；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11 and Z13 be III level, imprinted gene Z1, Z3, Z4, Z5, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z6, Z8, Z10, Z11 and Z13 is III level or marking base Imprinted gene loss of expression amount because being no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is IV grades And the imprinted gene copy number no more than 1 imprinted gene in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 Unconventionality expression amount is any one situation in IV grades, then is intermediate gastric carcinoma；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, In Z11 and Z13 the imprinted gene loss of expression amount of at least two imprinted gene be IV grades or imprinted gene Z1, Z3, Z4, Z5, Z6, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene is IV grades in Z8, Z10, Z11 and Z13, then is advanced gastric Cancer.

9. any in a kind of imprinted gene hierarchy model according to any one of claims 1 to 5 and/or such as claim 6-8 Device described in is used to prepare the drug or kit of esophageal tumor and/or stomach neoplasm detection and/or treatment.

10. purposes according to claim 9, which is characterized in that it is benign swollen to judge that the degree of benign and malignant of esophageal tumor is divided into Tumor, cancer of the esophagus potential, early esophageal cancer, mid-term cancer of the esophagus and advanced esophageal cancer；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount and imprinted gene copy number unconventionality expression amount of Z11, Z13 and Z16 are respectively less than I grades, imprinted gene The imprinted gene loss of expression amount no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is The marking base no more than 1 imprinted gene in I grades and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 It is then benign tumour because copy number unconventionality expression amount is any one situation in I grades；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11, Z13 and Z16 be I grades, imprinted gene Z1, Z3, Z4, Z5, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z6, Z8, Z10, Z11, Z13 and Z16 is I grades or the marking It is no more than the imprinted gene loss of expression amount of 1 imprinted gene in gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 For the marking no more than 1 imprinted gene in II grades and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 Gene copy number unconventionality expression amount is any one situation in II grades, then is cancer of the esophagus potential；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11, Z13 and Z16 be II grades, imprinted gene Z1, Z3, Z4, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z5, Z6, Z8, Z10, Z11, Z13 and Z16 be II grades or The imprinted gene for being no more than 1 imprinted gene in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 lacks table It is to be no more than 1 imprinted gene in III level and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 up to amount Imprinted gene copy number unconventionality expression amount be III level in any one situation, then be early esophageal cancer；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11, Z13 and Z16 be III level, imprinted gene Z1, Z3, Z4, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z5, Z6, Z8, Z10, Z11, Z13 and Z16 be III level or The imprinted gene for being no more than 1 imprinted gene in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 lacks table Up to amount for IV grade and in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 no more than 1 imprinted gene Imprinted gene copy number unconventionality expression amount is any one situation in IV grades, then is mid-term cancer of the esophagus；

Preferably, the result of the degree of benign and malignant for judging esophageal tumor for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, In Z11, Z13 and Z16 the imprinted gene loss of expression amount of at least two imprinted gene be IV grades or imprinted gene Z1, Z3, Z4, Z5, In Z6, Z8, Z10, Z11, Z13 and Z16 the imprinted gene copy number unconventionality expression amount of at least two imprinted gene be IV grades, then for Advanced esophageal cancer；

Preferably, the degree of benign and malignant of the judgement stomach neoplasm is divided into benign stomach neoplasm, gastric cancer potential, early carcinoma of stomach, mid-term stomach Cancer and late gastric cancer；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount and imprinted gene copy number unconventionality expression amount of Z11, Z13 and Z16 are respectively less than I grades, imprinted gene The imprinted gene loss of expression amount no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 is The marking base no more than 1 imprinted gene in I grades and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11, Z13 and Z16 It is then benign tumour because copy number unconventionality expression amount is any one situation in I grades；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11 and Z13 be I grades, imprinted gene Z1, Z3, Z4, Z5, Z6, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z8, Z10, Z11 and Z13 is I grades, imprinted gene Z1, The imprinted gene loss of expression amount no more than 1 imprinted gene in Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is I grades and prints Remember that the imprinted gene copy number for being no more than 1 imprinted gene in gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is abnormal Expression quantity be I grades and the imprinted gene loss of expression amount of imprinted gene Z16 or imprinted gene copy number unconventionality expression amount be I grades or More than or imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 in be no more than 1 imprinted gene imprinted gene lack Losing expression quantity is no more than 1 imprinted gene in II grade and imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 Imprinted gene copy number unconventionality expression amount is any one situation in II grades, then is gastric cancer potential；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11 and Z13 be II grades, imprinted gene Z1, Z3, Z4, Z5, Z6, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene in Z8, Z10, Z11 and Z13 is II grades or imprinted gene The imprinted gene loss of expression amount for being no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is III level And the imprinted gene copy number no more than 1 imprinted gene in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 Unconventionality expression amount is any one situation in III level, then is early carcinoma of stomach；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, The imprinted gene loss of expression amount of at least two imprinted gene in Z11 and Z13 be III level, imprinted gene Z1, Z3, Z4, Z5, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene of Z6, Z8, Z10, Z11 and Z13 is III level or marking base Imprinted gene loss of expression amount because being no more than 1 imprinted gene in Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 is IV grades And the imprinted gene copy number no more than 1 imprinted gene in imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, Z11 and Z13 Unconventionality expression amount is any one situation in IV grades, then is intermediate gastric carcinoma；

Preferably, the result of the degree of benign and malignant for judging stomach neoplasm for imprinted gene Z1, Z3, Z4, Z5, Z6, Z8, Z10, In Z11 and Z13 the imprinted gene loss of expression amount of at least two imprinted gene be IV grades or imprinted gene Z1, Z3, Z4, Z5, Z6, The imprinted gene copy number unconventionality expression amount of at least two imprinted gene is IV grades in Z8, Z10, Z11 and Z13, then is advanced gastric Cancer；

Preferably, a kind of marker of late gastric cancer diagnosis, which is characterized in that the marker is imprinted gene Z1 and marking base Because of Z11；

Wherein, the imprinted gene Z1 is Gnas, and the imprinted gene Z11 is Grb10；

Preferably, the late gastric cancer is that the TNM grade of gastric cancer is greater than the gastric cancer of T2N0M0.