CN103865996A - Endometrial cancer gene detection kit - Google Patents
Endometrial cancer gene detection kit Download PDFInfo
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- CN103865996A CN103865996A CN201310482915.6A CN201310482915A CN103865996A CN 103865996 A CN103865996 A CN 103865996A CN 201310482915 A CN201310482915 A CN 201310482915A CN 103865996 A CN103865996 A CN 103865996A
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- endometrium
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- 206010014759 Endometrial neoplasm Diseases 0.000 title claims abstract description 34
- 238000001514 detection method Methods 0.000 title abstract description 12
- 206010014733 Endometrial cancer Diseases 0.000 title abstract description 7
- 108700019961 Neoplasm Genes Proteins 0.000 title 1
- 102000048850 Neoplasm Genes Human genes 0.000 title 1
- 230000002357 endometrial effect Effects 0.000 claims abstract description 6
- 102000014160 PTEN Phosphohydrolase Human genes 0.000 claims abstract description 5
- 108010011536 PTEN Phosphohydrolase Proteins 0.000 claims abstract description 5
- 101150073900 PTEN gene Proteins 0.000 claims abstract 2
- 201000003914 endometrial carcinoma Diseases 0.000 claims description 29
- 108090000623 proteins and genes Proteins 0.000 claims description 24
- 238000012360 testing method Methods 0.000 claims description 14
- 210000004696 endometrium Anatomy 0.000 claims description 8
- 239000003153 chemical reaction reagent Substances 0.000 claims description 6
- 238000003745 diagnosis Methods 0.000 claims description 4
- 101150068332 KIT gene Proteins 0.000 claims 1
- 238000011065 in-situ storage Methods 0.000 claims 1
- 239000000523 sample Substances 0.000 abstract description 25
- 230000035772 mutation Effects 0.000 abstract description 3
- 108091028043 Nucleic acid sequence Proteins 0.000 abstract description 2
- 230000004913 activation Effects 0.000 abstract description 2
- 238000003752 polymerase chain reaction Methods 0.000 description 13
- 238000000034 method Methods 0.000 description 10
- 230000003321 amplification Effects 0.000 description 5
- 210000004027 cell Anatomy 0.000 description 5
- 238000003199 nucleic acid amplification method Methods 0.000 description 5
- 230000008569 process Effects 0.000 description 5
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 description 4
- 239000012634 fragment Substances 0.000 description 4
- 230000007170 pathology Effects 0.000 description 4
- 108020004414 DNA Proteins 0.000 description 3
- 206010028980 Neoplasm Diseases 0.000 description 3
- 201000011510 cancer Diseases 0.000 description 3
- 238000002405 diagnostic procedure Methods 0.000 description 3
- 201000010099 disease Diseases 0.000 description 3
- 239000000203 mixture Substances 0.000 description 3
- NBIIXXVUZAFLBC-UHFFFAOYSA-N Phosphoric acid Chemical compound OP(O)(O)=O NBIIXXVUZAFLBC-UHFFFAOYSA-N 0.000 description 2
- 230000008859 change Effects 0.000 description 2
- 230000000875 corresponding effect Effects 0.000 description 2
- 238000013016 damping Methods 0.000 description 2
- 239000003814 drug Substances 0.000 description 2
- 238000005516 engineering process Methods 0.000 description 2
- 238000000605 extraction Methods 0.000 description 2
- 239000012530 fluid Substances 0.000 description 2
- 238000000338 in vitro Methods 0.000 description 2
- 238000007689 inspection Methods 0.000 description 2
- 239000006210 lotion Substances 0.000 description 2
- 230000004060 metabolic process Effects 0.000 description 2
- 230000000869 mutational effect Effects 0.000 description 2
- 239000012188 paraffin wax Substances 0.000 description 2
- 238000004321 preservation Methods 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 238000012163 sequencing technique Methods 0.000 description 2
- 208000024891 symptom Diseases 0.000 description 2
- 210000004291 uterus Anatomy 0.000 description 2
- 238000005406 washing Methods 0.000 description 2
- 201000009030 Carcinoma Diseases 0.000 description 1
- 206010008342 Cervix carcinoma Diseases 0.000 description 1
- 238000007400 DNA extraction Methods 0.000 description 1
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 description 1
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 description 1
- 102000004190 Enzymes Human genes 0.000 description 1
- 108090000790 Enzymes Proteins 0.000 description 1
- 238000012408 PCR amplification Methods 0.000 description 1
- 102000052575 Proto-Oncogene Human genes 0.000 description 1
- 108700020978 Proto-Oncogene Proteins 0.000 description 1
- -1 Taq E Substances 0.000 description 1
- 102000044209 Tumor Suppressor Genes Human genes 0.000 description 1
- 108700025716 Tumor Suppressor Genes Proteins 0.000 description 1
- 208000006105 Uterine Cervical Neoplasms Diseases 0.000 description 1
- 229910000147 aluminium phosphate Inorganic materials 0.000 description 1
- 150000001413 amino acids Chemical class 0.000 description 1
- 230000008901 benefit Effects 0.000 description 1
- 230000000740 bleeding effect Effects 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- 201000010881 cervical cancer Diseases 0.000 description 1
- 238000004891 communication Methods 0.000 description 1
- 230000000295 complement effect Effects 0.000 description 1
- 230000002596 correlated effect Effects 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 239000012154 double-distilled water Substances 0.000 description 1
- 210000005168 endometrial cell Anatomy 0.000 description 1
- 210000004996 female reproductive system Anatomy 0.000 description 1
- 238000002657 hormone replacement therapy Methods 0.000 description 1
- 230000002779 inactivation Effects 0.000 description 1
- 230000001788 irregular Effects 0.000 description 1
- 230000003902 lesion Effects 0.000 description 1
- 239000007788 liquid Substances 0.000 description 1
- 206010025482 malaise Diseases 0.000 description 1
- 239000003550 marker Substances 0.000 description 1
- 230000035800 maturation Effects 0.000 description 1
- 210000000214 mouth Anatomy 0.000 description 1
- 230000008506 pathogenesis Effects 0.000 description 1
- 210000005259 peripheral blood Anatomy 0.000 description 1
- 239000011886 peripheral blood Substances 0.000 description 1
- 208000030683 polygenic disease Diseases 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- 230000001105 regulatory effect Effects 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 230000004044 response Effects 0.000 description 1
- 238000012216 screening Methods 0.000 description 1
- 230000004083 survival effect Effects 0.000 description 1
- 238000012795 verification Methods 0.000 description 1
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 1
Classifications
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- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6876—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
- C12Q1/6883—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material
- C12Q1/6886—Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes for diseases caused by alterations of genetic material for cancer
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q2600/00—Oligonucleotides characterized by their use
- C12Q2600/156—Polymorphic or mutational markers
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- Chemical & Material Sciences (AREA)
- Life Sciences & Earth Sciences (AREA)
- Health & Medical Sciences (AREA)
- Organic Chemistry (AREA)
- Proteomics, Peptides & Aminoacids (AREA)
- Engineering & Computer Science (AREA)
- Immunology (AREA)
- Pathology (AREA)
- Analytical Chemistry (AREA)
- Zoology (AREA)
- Genetics & Genomics (AREA)
- Wood Science & Technology (AREA)
- Physics & Mathematics (AREA)
- Biotechnology (AREA)
- Microbiology (AREA)
- Molecular Biology (AREA)
- Hospice & Palliative Care (AREA)
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- Oncology (AREA)
- Biochemistry (AREA)
- Bioinformatics & Cheminformatics (AREA)
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Abstract
The invention discloses a kit for endometrial cancer detection for an endometrial tissue sample. The detection is based on the activation mutation of a specific site in a PTEN gene. The invention relates to an endometrial cancer detection kit and probe for endometrial cancer detection based on related nucleotide sequences of the PTEN mutation site.
Description
Technical field
The present invention is for to uterine endometrium tissue sample, relates to application and relates to related nucleotide sequences, primer and the probe based on PTEN mutational site, the carcinoma of endometrium detection kit that carcinoma of endometrium is detected.
Background technology
Carcinoma of endometrium is primary in uterine endometrium, is one of women's three large common cancers.Endometrial malignant neoplasms has exceeded cervical cancer closely over several years becomes the modal aggressive malignant tumour of female reproductive system.General this disease only occurs women just implementing to check when the symptoms such as irregular bleeding are carried out the inspection of relative disease.Thereby cause most because being difficult for finding without the patient of clinical symptom in early days in pathology, once and be found, the more late survival rate of its staging diagnosis is lower, therefore in asymptomatic crowd, carries out examination significant to find precancerous lesion and early-stage cancer.
The examination of carcinoma of endometrium disease in the past often needs to examine under a microscope ten hundreds of tissue slicies, and cell interior composition, cellular form and behavior, cell colony characteristic etc. are analyzed.For existing issue, my company is engaged in carcinoma of endometrium screening and recognizes that with medical worker's communication carcinoma of endometrium is a class polygenic disease based on coordinating with medical institutions for many years, its evolution complexity, clinical manifestation is various, relate to the variation of multiple genes relevant with many factors, due to the widespread use of hormone replacement therapy, sickness rate rises year by year, but its pathogenesis is illustrated not yet completely.Research shows, the generation of carcinoma of endometrium is relevant with the inactivation of the activation of some proto-oncogene and antioncogene.Clinical medicine needs a kind of simple and easy to do endometrial cell tissue to collecting to carry out rapid gene diagnostic method.
The relevant patent detecting for carcinoma of endometrium has various ways at present.As protein expression water product: " detection of carcinoma of endometrium " publication number: CN1257550A, " carcinoma of endometrium diagnostic reagent, test kit and control medicine " publication number: CN101261279A, CN101261279B; Undertaken or oral cavity sample detects by uterus washing lotion, as " marker of carcinoma of endometrium " publication number: CN102549169A, invention is detected in " Non-invasive detection kit for endometrial carcinoma predisposing gene " publication number: CN102559903A and other gene test sites, and related gene locus is different from the site the present invention relates to.
Multiple invention above, because other site samples are through Multiple Cycle in human body, may be more because relating to process, result is judged to the error causing; And the pathogeny of current carcinoma of endometrium is not clear and definite, need the higher detection method of a kind of accuracy, what the carcinoma of endometrium making can be more early stage is detected.
Summary of the invention
Object of the present invention: in order to research and develop a kind of endometrial tissue sample that can directly collect for medical institutions, carry out the inspection of carcinoma of endometrium, go out sample donor with early detection and whether suffer from carcinoma of endometrium.Potential endometrial carcinoma is effectively made a definite diagnosis in time, given treatment to.
Specific embodiments of the present invention: the present invention adopts molecular biological diagnostic method, obtain and polymerase chain amplification (PCR) are carried out in corresponding site after sample and obtain after a large amount of products, check order, and analytical results.Composition comprises: the explanation of diagnostic reagent+using method
Integral part one diagnostic reagent of the present invention: comprise the required enzyme of PCR (polymerase chain reaction), damping fluid, target primer and correlated response system.PCR (polymerase chain reaction) is the replication in vitro process of DNA, utilize DNA 95 to untwist while spending in vitro, 35 primers while spending and strand match combination the more synthetic complementary strand of the direction of temperature regulating to 65 degree left and right archaeal dna polymerase along phosphoric acid to five-carbon sugar (5 '-3 ') by base complementrity.Obtain a large amount of target genes.
Integral part of the present invention-using method explanation: the pretreatment process, the gene extracting method parameter that comprise the endometrium cast-off cells kind sample form that paraffin specimen, preservation liquid are preserved; The parameters such as PCR (polymerase chain reaction) time, temperature; Judge the sequencing result foundation of target gene.
Experimental verification premenstruum, deep comparison discovery PTEN c.946C this point mutation of > T does not cause amino acid whose change, but in Endometrial Carcinomas group, recall rate is up to 83%, in film propagation group, do not find the sudden change of the sample of any degree in uterus, specificity, up to 100%, can be used for pointing out the generation of carcinoma of endometrium.
Target gene site is chosen: PTENNM_000314.4c.946C > T is carcinoma of endometrium examination site.
The specific variations in reply target gene site, is used the primer fragment for its development specially, and target fragment is increased in a large number, is convenient to late detection.
The a large amount of target fragment that obtain by the PCR to sample gene (polymerase chain reaction) are carried out gene sequencing, judge that whether its gene order is identical with standard sequence.
[0024] the present invention and similar similar comparing, the present invention directly adopts uterine endometrium sample as detection resources, directly chooses the tissue sample of position of disease, make detected result still less receive interference.As take other sample modes, for example uterine cavity washing lotion, peripheral blood, may, because of the factor such as pathology, organism metabolism at other positions of body, be interfered detected result, can not specify pathology kind or make to fail to detect pathology because metabolism changes.
The method of the sample gene extraction that uses in the Realization of Product of place of the present invention, pcr amplification, gene test, process, equipment etc. all have certain basis at home, and level of automation is higher, technology maturation.
Sample required for the present invention is a small amount of, and the erasion of examining that both can choose medical institutions obtains sample, and the micro-sample that also can use the endometrium collector tool of the micro-damage using in gynecologial examination to take detects.Because of its sample that can use the gynecologial examination of medical institutions, do not need to increase more process, directly the inner membrance sample of its collection is detected, more carcinoma of endometrium is detected in time.
The test kit detecting with respect to other carcinomas of endometrium, adopts endometrial tissue sample to carry out gene test, has more advantage on interference free performance.In actual use, there are multiple medical examination means to take uterine endometrium sample, this test kit can be applied.As fractional curettage, uterine cell collector etc.
In sum, use the specific mutational site of PTEN to can be used as carcinoma of endometrium gene test position, and practical application other are smoothly relevant, so application this patent.
Embodiment
Test kit composition: PCR damping fluid, ddH2O, Taq E, primer and dNTP
The purposes of test kit: this test kit is take endometrial slough tissue cell, paraffin specimen as detected object, specific objective gene in sample is increased and checked order by polymerase chain amplification (PCR) method, to examination endometrial carcinoma from carcinoma of endometrium group of people at high risk
Actual in test kit should be left-20 ℃ in, be kept in Dark Place, and validity period is 1 year.Often use after using for the first time and be positioned over-4 ℃ of preservations, effective in March.
Test kit principle summary: this product adopts molecular biological diagnostic method, obtains and polymerase chain amplification (PCR) is carried out in corresponding site after sample and obtain after a large amount of products, checks order, and analytical results.
Sample is carried out to DNA extraction.
The target dna of extraction is carried out to polymerase chain amplification (PCR).
By the gene fragment order-checking after amplification, contrast with standard sequence.
Result of determination.
Sequence table
Claims (3)
1. carry out carcinoma of endometrium and carry out a gene diagnosis kit, contain the gene diagnosis reagent that uterine endometrium tissue in situ sample is gone out to carcinoma of endometrium.This test kit is the sequence of PTEN gene in assessment sample.As PTEN c.946C > T illustrate to there is carcinoma of endometrium feature.
2. diagnostic reagent according to claim 1, is characterized in that: the check point of judging carcinoma of endometrium is SEQ ID NO:88 site, and the primer designing based on this site is realized the gene test means that carcinoma of endometrium is detected.
3. diagnostic reagent according to claim 1, is characterized in that: the kit gene that endometrial tissue is detected.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310482915.6A CN103865996A (en) | 2013-10-16 | 2013-10-16 | Endometrial cancer gene detection kit |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN201310482915.6A CN103865996A (en) | 2013-10-16 | 2013-10-16 | Endometrial cancer gene detection kit |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN103865996A true CN103865996A (en) | 2014-06-18 |
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Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN201310482915.6A Pending CN103865996A (en) | 2013-10-16 | 2013-10-16 | Endometrial cancer gene detection kit |
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| CN (1) | CN103865996A (en) |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110373469A (en) * | 2019-08-02 | 2019-10-25 | 重庆大学附属肿瘤医院 | A kind of capture sequencing probe of carcinoma of endometrium personalized medicine and preparation method thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN101261279A (en) * | 2008-04-11 | 2008-09-10 | 四川大学 | Endometrial cancer diagnostic reagents, kits and prevention and treatment drugs |
| CA2733672A1 (en) * | 2007-08-16 | 2009-02-19 | The Royal Institution For The Advancement Of Learning/Mcgill University | Tumor cell-derived microvesicles |
| CN102559903A (en) * | 2012-02-01 | 2012-07-11 | 解码（上海）生物医药科技有限公司 | Non-invasive detection kit for endometrial carcinoma predisposing gene |
-
2013
- 2013-10-16 CN CN201310482915.6A patent/CN103865996A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CA2733672A1 (en) * | 2007-08-16 | 2009-02-19 | The Royal Institution For The Advancement Of Learning/Mcgill University | Tumor cell-derived microvesicles |
| CN101261279A (en) * | 2008-04-11 | 2008-09-10 | 四川大学 | Endometrial cancer diagnostic reagents, kits and prevention and treatment drugs |
| CN102559903A (en) * | 2012-02-01 | 2012-07-11 | 解码（上海）生物医药科技有限公司 | Non-invasive detection kit for endometrial carcinoma predisposing gene |
Non-Patent Citations (1)
| Title |
|---|
| 张祖娟: "PTEN基因突变在子宫内膜癌早期诊断中的意义及价值", 《中国优秀硕士学位论文全文数据库医药卫生科技辑》 * |
Cited By (1)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN110373469A (en) * | 2019-08-02 | 2019-10-25 | 重庆大学附属肿瘤医院 | A kind of capture sequencing probe of carcinoma of endometrium personalized medicine and preparation method thereof |
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