CN108866189B - A kit and system for predicting susceptibility to laryngeal squamous cell carcinoma - Google Patents

Abstract

The present invention relates to a kit for predicting susceptibility to laryngeal squamous cell carcinoma, comprising the following components: STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, STR-6 primer Primer; further, it can also include: PCR amplification reaction solution, LIZ-500 molecular weight internal standard, deionized formamide. The laryngeal squamous cell carcinoma susceptibility prediction kit of the present invention can be used for the diagnosis and susceptibility prediction of laryngeal squamous cell carcinoma. The present invention also provides a susceptibility prediction system for laryngeal squamous cell carcinoma.

Description

A kit and system for predicting susceptibility to laryngeal squamous cell carcinoma

technical field

The present invention relates to the field of biomedicine. Specifically, it relates to a laryngeal squamous cell carcinoma susceptibility prediction kit and a laryngeal squamous cell carcinoma susceptibility prediction system. More specifically, the present invention relates to a kit for detecting STR of laryngeal squamous cell carcinoma susceptibility-related genes by a method for analyzing short tandem repeats (Shorttandem repeats, referred to as STR) site fragments, and by combining discriminant analysis statistical methods, In this way, an early warning of the susceptibility of laryngeal squamous cell carcinoma in the subject is carried out.

Background technique

Tumor is a disease closely related to genetics. To study the molecular genetic basis of tumor, and then propose tumor-specific genetic markers, it is hoped that it can be used for routine detection, clinical diagnosis, personalized treatment, and follow-up. etc. to provide a simple and feasible method. However, the individual differences of patients and the intersection of related biomolecular events at different developmental stages have brought great difficulties to this work.

Laryngeal squamous cell carcinoma is the most common malignant tumor of larynx. When the tumor enlarges and the surface ulcerates, it can cause swallowing pain, and ipsilateral reflex earache, often accompanied by progressive dysphagia. Tumors involving the larynx can cause difficulty breathing and hoarseness. The incidence of laryngeal squamous cell carcinoma is affected by certain living habits and other external factors. The incidence of males is higher, and it is related to living habits such as smoking and alcoholism. Therefore, predicting the susceptibility of laryngeal squamous cell carcinoma in the examined population is helpful to improve the risk awareness of the disease. The prediction results show that people with a higher risk of laryngeal squamous cell carcinoma can live by quitting smoking and drinking. Habits to reduce the risk of disease or early detection and early treatment.

A large number of studies have shown that genetic polymorphisms of tumor-related genes play a key role in the occurrence and development of malignant tumors. However, the occurrence and development of tumors is a very complex process, and it is obviously impossible and unscientific to use the changes of a single molecular genetic marker to diagnose the disease. With the existing technical means, it is still not possible to carry out a relatively accurate early warning of tumor susceptibility only through genetic information, and the current early identification and prediction methods for tumors still need to be improved. The invention relates to a reagent for early warning of the susceptibility of laryngeal squamous cell carcinoma by combined detection of multiple STR loci with high correlation with the occurrence of laryngeal squamous cell carcinoma by means of STR locus fragment analysis, combined with a discriminant analysis and statistical method box.

SUMMARY OF THE INVENTION

In order to solve the above-mentioned problems existing in the prior art, the present invention relates to the joint detection of multiple STR loci with high correlation with the occurrence of laryngeal squamous cell carcinoma through the STR locus fragment analysis method, combined with the discriminant analysis statistical method, and the detection of laryngeal squamous cell carcinoma. Squamous cell carcinoma susceptibility for early warning.

The present invention is based on the following findings of the inventors: the inventors analyzed the STR of the genomic DNA of the laryngeal squamous cell carcinoma subjects and the healthy control subjects, and found that in a large number of laryngeal squamous cell carcinoma samples and control samples After verification, it was found that the number of short tandem sequence repeats at each independent STR locus was not significantly associated with laryngeal squamous cell carcinoma, while the combination of the number of short tandem sequence repeats at some specific STR loci was significantly correlated with the subject. Suffering from laryngeal squamous cell carcinoma is closely related.

To this end, the present invention proposes an isolated set of STR loci that are highly associated with developing laryngeal squamous cell carcinoma. According to embodiments of the present invention, these isolated STR loci comprise the nucleotide sequences shown in STR-1 to STR-6 (Table 1). Using these isolated STR loci as a reference, the susceptibility of laryngeal squamous cell carcinoma can be effectively predicted.

Table 1

Site code starting point gene short tandem sequence STR-1 X chromosome, position 66657655 AR CAG STR-2 Chromosome 4, position 55633758 Bat-25 T STR-3 Chromosome 5, position 111646983 D5S346 GT STR-4 Chromosome 6, position 151806531 ER1 TA STR-5 Chromosome 14, position 64253561 ER2 TG STR-6 Chromosome 4, position 154587748 FGA AAAG

For the detailed description of the above-mentioned STR sites, those skilled in the art can log in to relevant databases (such as GeneBank, Nucleotide, etc.) to obtain, and will not be repeated here. The inventor surprisingly found that by analyzing the cell genome of the subject to obtain the repetitions of the short tandem sequence of each STR locus above, and using the repetitions as an independent variable to perform statistical analysis methods such as discriminant analysis, it is possible to detect laryngeal squamous cells. Cell cancer susceptibility for early warning.

On this basis, one of the technical problems solved by the present invention is to provide a kit for predicting susceptibility to laryngeal squamous cell carcinoma, which includes the following components: STR-1 primer, STR-2 primer, STR-3 primer , STR-4 primer, STR-5 primer, STR-6 primer, the above primers are respectively used to amplify the target fragment containing the short tandem sequences listed in Table 1, so as to determine the number of repetitions of the short tandem sequences.

Preferably, the laryngeal squamous cell carcinoma susceptibility prediction kit of the present invention further comprises: PCR amplification reaction solution, LIZ-500 molecular weight internal standard, and deionized formamide.

In the laryngeal squamous cell carcinoma susceptibility prediction kit of the present invention, preferably, the sequences of the above STR-1 primers, STR-2 primers, STR-3 primers, STR-4 primers, STR-5 primers, and STR-6 primers As shown in Table 2 below, more preferably, the concentration of each of the above primers is 10 μM:

Table 2

In Table 2, HEX, FAM, and ROX are all fluorescent groups for labeling the 5' end, HEX is hexachloro-6-methylfluorescein, FAM is 6-carboxyfluorescein, and ROX is ROX reference dye.

In the laryngeal squamous cell carcinoma susceptibility prediction kit of the present invention, preferably, the PCR amplification reaction solution is a mixture of the following reagents: TaqDNA polymerase (5 U/μL), Tris-HCl (100 mM, at 25° C. pH 8.8), KCl (500 mM), ethylphenyl polyethylene glycol (0.8% (v/v)), _MgCl2 (25 mM), dNTPs (10 mM), deionized water.

More preferably, the PCR amplification reaction solution is stored at -20°C.

In the laryngeal squamous cell carcinoma susceptibility prediction kit of the present invention, preferably, the LIZ-500 molecular weight internal standard can be stored at -20°C;

In the laryngeal squamous cell carcinoma susceptibility prediction kit of the present invention, preferably, the deionized formamide can be stored at 2-8°C.

Preferably, the laryngeal squamous cell carcinoma susceptibility prediction kit of the present invention further comprises an instruction manual.

The instruction manual describes the use method of the above-mentioned laryngeal squamous cell carcinoma susceptibility prediction kit, which comprises the following steps:

(1) Extract sample DNA;

(2) PCR reaction

(2-1) Take out the STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, STR-6 primer, and PCR amplification reaction solution from the refrigerator, and equilibrate to room temperature. After the components are fully dissolved, centrifuge quickly for 10 seconds;

(2-2) Take 30-300ng of sample DNA, add 60μL of PCR amplification reaction solution, add deionized water to make up to 115.2μL, mix well, centrifuge quickly for 10 seconds, and then dispense the mixture at 19.2μL/well to 6 PCR reaction tubes;

(2-3) Add STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, and STR-6 primer to step (2-2) at 0.8 μL/well respectively. 6 PCR reaction tubes; cover the PCR reaction tube caps, record the sample loading situation, centrifuge quickly for 10 seconds, then transfer the PCR reaction tubes to the corresponding position of the sample tank of the PCR amplifier, record the placement sequence, and start PCR amplification Amplification reaction conditions: 95°C for 3 minutes; 95°C for 30 seconds, 60°C for 30 seconds, 72°C for 30 seconds, 10 cycles; 95°C for 30 seconds, 55°C for 30 seconds, 72°C for 30 seconds, 20 cycles ; 6 minutes at 72°C to obtain 6 sets of PCR amplification products;

(3) STR fragment analysis

(3-1) Take 990 μL of deionized formamide, add 10 μL of LIZ-500 molecular weight internal standard, mix well, centrifuge quickly for 10 seconds, add 10 μL/well to sequencing reaction tubes, and centrifuge quickly for 10 seconds;

(3-2) Add 1 μL/well of the above-mentioned 6 sets of PCR amplification products to 6 sequencing reaction tubes, and centrifuge quickly for 10 seconds; then transfer the sequencing reaction tubes to the corresponding position of the sample tank of the PCR amplifier, 98°C Heating for 5 minutes. Immediately after the end of the program, the sequencing reaction tube was placed on the ice-water mixture and rapidly cooled to 0 °C, and centrifuged rapidly for 10 seconds; then the sequencing reaction tube was transferred to the corresponding position of the sample tank of the STR locus fragment analyzer, and recorded and placed sequence, perform fragment analysis and detection;

(4) Analysis and judgment of results

(4-1) According to the fragment analysis results, record the fragment lengths of the two alleles of STR-1, STR-2, STR-3, STR-4, STR-5, and STR-6 respectively:

The smaller fragment length value of the two alleles of STR-1 is recorded as L ₁ , and the larger fragment length value of the two alleles of STR-1 is recorded as L ₂ ;

The smaller fragment length value of the two alleles of STR-2 is recorded as L ₃ , and the larger fragment length value of the two alleles of STR-2 is recorded as L ₄ ;

The smaller fragment length value of the two alleles of STR-3 is recorded as L ₅ , and the larger fragment length value of the two alleles of STR-3 is recorded as L ₆ ;

The smaller fragment length value of the two alleles of STR-4 is recorded as L ₇ , and the larger fragment length value of the two alleles of STR-4 is recorded as L ₈ ;

The smaller fragment length value of the two alleles of STR-5 is recorded as L ₉ , and the larger fragment length value of the two alleles of STR-5 is recorded as L ₁₀ ;

The smaller fragment length value of the two alleles of STR-6 is recorded as L ₁₁ , and the larger fragment length value of the two alleles of STR-6 is recorded as L ₁₂ ;

(4-2) The number of repetitions of the short tandem sequence is calculated according to the fragment length and the following formula, and is denoted as X ₁ -X ₁₂ , where round represents rounding to an integer:

X ₁ =round[(L ₁ -191)/3]; X ₂ =round[(L ₂ -191)/3];

X ₃ =round(L ₃ -379); X ₄ =round(L ₄ -379);

X ₅ =round[(L ₅ -202)/2]; X ₆ =round[(L ₆ -202)/2];

X ₇ =round[(L ₇ -359)/2]; X ₈ =round[(L ₈ -359)/2];

X ₉ =round[(L ₉ -278)/2]; X ₁₀ =round[(L ₁₀ -278)/2];

X ₁₁ =round[(L ₁₁ -200)/4]; X ₁₂ =round[(L ₁₂ -200)/4];

(4-3) Substitute the repetition times of the above short series sequence into the preset discriminant function:

F _LC = 10.750X ₁ -8.272X ₂ +14.088X ₃ +106.284X ₄ -0.418X ₅ +2.056X ₆ +2.288X ₇ +

1.131X ₈ -0.466X ₉ -4.985X ₁₀ +2.407X ₁₁ +7.072X ₁₂ -15.282X ₁₃ -1550.869

F _LN = 11.048X ₁ -8.564X ₂ +14.325X ₃ +108.309X ₄ -0.776X ₅ +1.944X ₆ +2.347X ₇ +

0.795X ₈ -0.807X ₉ -4.929X ₁₀ +2.523X ₁₁ +6.583X ₁₂ -18.780X ₁₃ -1581.097

Wherein, if the subject is female, X ₁₃ =0; if the subject is male, X ₁₃ =1;

(4-4) Prediction of susceptibility to laryngeal squamous cell carcinoma:

Comparing the F _LC value and the F _LN value, if F _LC > F _LN , the probability of the subject suffering from laryngeal squamous cell carcinoma is predicted to be ≥82.4%; if F _LC ≤ F _LN , it is predicted that the subject will not suffer from laryngeal squamous cell carcinoma The chance of cell carcinoma is ≥83.3%.

In the present invention,

Preferably, the extraction of sample DNA in step (1) can use a purchased commercial genomic DNA extraction kit and operate according to the kit instructions. The sample can be whole blood, buccal swab or oral tissue of the subject, preferably whole blood of the subject.

Preferably, the rotational speed of all centrifugations in the method of use is preferably 3000 g/min.

The probability of suffering from laryngeal squamous cell carcinoma mentioned in the present invention is the sum of the probability of having suffered from laryngeal squamous cell carcinoma and the probability of suffering from laryngeal squamous cell carcinoma in the future. Therefore, it can be used not only for the diagnosis of laryngeal squamous cell carcinoma, but also for the risk warning of future laryngeal squamous cell carcinoma. and other ways to reduce the risk of laryngeal squamous cell carcinoma.

The second technical problem solved by the present invention is to provide a method for predicting the susceptibility of laryngeal squamous cell carcinoma, that is, using the above-mentioned kit and operating according to the method described in the specification.

The third technical problem solved by the present invention is to provide the application of the laryngeal squamous cell carcinoma susceptibility prediction kit in the preparation of laryngeal squamous cell carcinoma diagnostic products.

The fourth technical problem solved by the present invention is to provide a susceptibility prediction system for laryngeal squamous cell carcinoma, which includes:

A device for obtaining the number of repetitions of the short tandem sequence of the STR site of the sample DNA;

A data processing and determination device, which includes the following modules:

The data input module is used to input the age, gender, and repetition times of the short tandem sequence of the STR locus;

The database management module is used for the operation management of data storage, modification, deletion, query and printing;

The data calculation module is used to calculate the discriminant function result according to the repetition times of the short tandem sequence of the STR site in the data input module;

The analysis discriminant and result output module is used to compare the discriminant function results, so as to predict the susceptibility of laryngeal squamous cell carcinoma, and output the result.

in,

The number of repetitions of the short tandem sequences at the STR site is the number of repetitions of the following 6 pairs of short tandem sequences at the STR site:

Site code starting point gene short tandem sequence STR-1 X chromosome, position 66657655 AR CAG STR-2 Chromosome 4, position 55633758 Bat-25 T STR-3 Chromosome 5, position 111646983 D5S346 GT STR-4 Chromosome 6, position 151806531 ER1 TA STR-5 Chromosome 14, position 64253561 ER2 TG STR-6 Chromosome 4, position 154587748 FGA AAAG

The discriminant function includes:

First discriminant function F _LC =10.750X ₁ -8.272X ₂ +14.088X ₃ +106.284X ₄ -0.418X ₅ +2.056X ₆ +2.288X ₇ +1.131X ₈ -0.466X ₉ -4.985X ₁₀ +2.407X ₁₁ +7.072X ₁₂ -15.282X ₁₃ -1550.869

Second discriminant function F _LN =11.048X ₁ -8.564X ₂ +14.325X ₃ +108.309X ₄ -0.776X ₅ +1.944X ₆ +2.347X ₇ +0.795X ₈ -0.807X ₉ -4.929X ₁₀ +2.523X ₁₁ +6.583X ₁₂ -18.780X ₁₃ -1581.097

In the discriminant function,

X ₁ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-1;

X ₂ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-1;

X ₃ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-2;

X ₄ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-2;

X ₅ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-3;

X ₆ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-3;

X ₇ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-4;

X ₈ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-4;

X ₉ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-5;

X ₁₀ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-5;

X ₁₁ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-6;

X ₁₂ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-6;

If the subject is female, X ₁₃ =0, and if the subject is male, X ₁₃ =1.

Among them, X ₁ ^- X ₁₂ are calculated according to the fragment length and the following formula, where round represents rounding to an integer:

X ₁ =round[(L ₁ -191)/3]; X ₂ =round[(L ₂ -191)/3];

X ₃ =round(L ₃ -379); X ₄ =round(L ₄ -379);

X ₅ =round[(L ₅ -202)/2]; X ₆ =round[(L ₆ -202)/2];

X ₇ =round[(L ₇ -359)/2]; X ₈ =round[(L ₈ -359)/2];

X ₉ =round[(L ₉ -278)/2]; X ₁₀ =round[(L ₁₀ -278)/2];

X ₁₁ =round[(L ₁₁ -200)/4]; X ₁₂ =round[(L ₁₂ -200)/4];

In X ₁ ^- X ₁₂ , L ₁ is the smaller fragment length value of the two alleles of STR-1, and L ₂ is the larger fragment length value of the two alleles of STR-1;

L ₃ is the smaller fragment length value of the two alleles of STR-2, and L ₄ is the larger fragment length value of the two alleles of STR-2;

L ₅ is the smaller fragment length value of the two alleles of STR-3, and L ₆ is the larger fragment length value of the two alleles of STR-3;

L ₇ is the smaller fragment length value of the two alleles of STR-4, and L ₈ is the larger fragment length value of the two alleles of STR-4;

L ₉ is the smaller fragment length value of the two alleles of STR-5, and L ₁₀ is the larger fragment length value of the two alleles of STR-5;

L ₁₁ is the smaller fragment length value of the two alleles of STR-6, and L ₁₂ is the larger fragment length value of the two alleles of STR-6;

The analysis, discrimination and result output module compares the calculation result of the first discriminant function F _LC with the calculation result of the second discriminant function F _LN , and if F _LC > F _LN , it outputs “The subject suffers from laryngeal squamous cells. If F _LC ≤ F _LN , output the prediction result of "the probability of the subject not suffering from laryngeal squamous cell carcinoma ≥ 83.3%".

The device for obtaining the repetition times of the short tandem sequence of the STR site of the sample DNA may include a STR site fragment analyzer, a PCR amplifier, etc.; the data processing and determination device may be a computer or other equipment.

The fifth technical problem solved by the present invention is to provide the application of the laryngeal squamous cell carcinoma susceptibility prediction system in the preparation of laryngeal squamous cell carcinoma prediction products, laryngeal squamous cell carcinoma diagnostic products, and oral and throat health auxiliary products.

The sixth technical problem to be solved by the present invention is to provide a laryngeal squamous cell carcinoma prediction product, a laryngeal squamous cell carcinoma diagnosis product, or an oral and throat health auxiliary product, which comprises the laryngeal squamous cell carcinoma Cancer Susceptibility Prediction System.

The test material used in the present invention is human genomic DNA. In theory, human genomic DNA will not change in a person's life. Human genomic DNA encodes all life activities of human beings, so theoretically, by detecting genomic DNA, the risk of a subject suffering from a certain disease can be predicted early, even at birth. The present invention is based on this theory, so using The kit and system of the present invention can not only play the role of early warning, but also play the role of diagnosis.

The occurrence and development of tumors is a very complex process. To study the molecular genetic basis of tumors, it is hoped that it can provide simple and feasible methods for routine detection, clinical diagnosis, personalized treatment, and follow-up of the disease after recovery. However, the individual differences of patients and the intersection of related biomolecular events at different developmental stages have brought great difficulties to this work. Using a single molecular genetic change to diagnose a tumor is clearly impossible and unscientific. The inventor uses modern molecular biology technology to jointly analyze multiple STRs in the genomic DNA of the subject, combined with statistical analysis methods such as discriminant analysis, thereby inventing a kit for early warning of laryngeal squamous cell carcinoma susceptibility .

Description of drawings

FIG. 1 is a schematic diagram of the modules included in the data processing and determination device in the laryngeal squamous cell carcinoma susceptibility prediction system according to the present invention.

Detailed ways

The specific embodiments of the present invention will be described below with reference to the accompanying drawings and examples, so as to better understand the present invention. The experimental methods used in the following examples are conventional methods unless otherwise specified. The following describes in detail the embodiments of the present invention, examples of which are illustrated in the accompanying drawings, wherein the same or similar reference numerals refer to the same or similar elements or elements having the same or similar functions throughout. The embodiments described below with reference to the accompanying drawings are exemplary, only used to explain the present invention, and should not be construed as a limitation of the present invention.

The PCR amplification instrument in the embodiment is the Mastercycler nexus amplification instrument (purchased from Eppendorf, USA);

The STR locus fragment analyzer in the examples is a 3730XL sequencing analyzer (purchased from ABI, USA);

The DNA extraction kit in the embodiment is blood DNAout kit (purchased from Beijing Tianenze Company);

The rotational speed of all centrifugation in the examples was 3000 g/min.

Example 1 A susceptibility prediction system for laryngeal squamous cell carcinoma

A laryngeal squamous cell carcinoma susceptibility prediction system, comprising:

A device for obtaining the number of repetitions of the short tandem sequence of the STR site of the sample DNA;

A data processing and determination device, which includes the following modules (as shown in Figure 1):

The data input module is used to input the age, gender, and repetition times of the short tandem sequence of the STR locus;

The database management module is used for the operation management of data storage, modification, deletion, query and printing;

The data calculation module is used to calculate the discriminant function result according to the repetition times of the short tandem sequence of the STR site in the data input module;

The analysis discriminant and result output module is used to compare the discriminant function results, so as to predict the susceptibility of laryngeal squamous cell carcinoma, and output the result.

in,

The number of repetitions of the short tandem sequences at the STR site is the number of repetitions of the following 6 pairs of short tandem sequences at the STR site:

The discriminant function includes:

First discriminant function F _LC =10.750X ₁ -8.272X ₂ +14.088X ₃ +106.284X ₄ -0.418X ₅ +2.056X ₆ +2.288X ₇ +1.131X ₈ -0.466X ₉ -4.985X ₁₀ +2.407X ₁₁ +7.072X ₁₂ -15.282X ₁₃ -1550.869

Second discriminant function F _LN =11.048X ₁ -8.564X ₂ +14.325X ₃ +108.309X ₄ -0.776X ₅ +1.944X ₆ +2.347X ₇ +0.795X ₈ -0.807X ₉ -4.929X ₁₀ +2.523X ₁₁ +6.583X ₁₂ -18.780X ₁₃ -1581.097

In the discriminant function,

X ₁ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-1;

X ₂ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-1;

X ₃ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-2;

X ₄ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-2;

X ₅ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-3;

X ₆ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-3;

X ₇ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-4;

X ₈ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-4;

X ₉ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-5;

X ₁₀ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-5;

X ₁₁ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-6;

X ₁₂ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-6;

If the subject is female, X ₁₃ =0, and if the subject is male, X ₁₃ =1.

Among them, X ₁ ^- X ₁₂ are calculated according to the fragment length and the following formula, where round represents rounding to an integer:

X ₁ =round[(L ₁ -191)/3]; X ₂ =round[(L ₂ -191)/3];

X ₃ =round(L ₃ -379); X ₄ =round(L ₄ -379);

X ₅ =round[(L ₅ -202)/2]; X ₆ =round[(L ₆ -202)/2];

X ₇ =round[(L ₇ -359)/2]; X ₈ =round[(L ₈ -359)/2];

X ₉ =round[(L ₉ -278)/2]; X ₁₀ =round[(L ₁₀ -278)/2];

X ₁₁ =round[(L ₁₁ -200)/4]; X ₁₂ =round[(L ₁₂ -200)/4];

In X ₁ ^- X ₁₂ , L ₁ is the smaller fragment length value of the two alleles of STR-1, and L ₂ is the larger fragment length value of the two alleles of STR-1;

L ₃ is the smaller fragment length value of the two alleles of STR-2, and L ₄ is the larger fragment length value of the two alleles of STR-2;

L ₅ is the smaller fragment length value of the two alleles of STR-3, and L ₆ is the larger fragment length value of the two alleles of STR-3;

L ₇ is the smaller fragment length value of the two alleles of STR-4, and L ₈ is the larger fragment length value of the two alleles of STR-4;

L ₉ is the smaller fragment length value of the two alleles of STR-5, and L ₁₀ is the larger fragment length value of the two alleles of STR-5;

L ₁₁ is the smaller fragment length value of the two alleles of STR-6, and L ₁₂ is the larger fragment length value of the two alleles of STR-6;

The analysis, discrimination and result output module compares the calculation result of the first discriminant function F _LC with the calculation result of the second discriminant function F _LN , and if F _LC > F _LN , it outputs “The subject suffers from laryngeal squamous cells. If F _LC ≤ F _LN , output the prediction result of “the probability that the subject does not suffer from laryngeal squamous cell carcinoma ≥ 83.3%”.

Example 2 A kit for predicting susceptibility to laryngeal squamous cell carcinoma

A kit for predicting susceptibility to laryngeal squamous cell carcinoma, comprising the following components: STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, STR-6 primer, PCR Amplification reaction solution, LIZ-500 molecular weight internal standard, deionized formamide, instruction manual.

The concentrations of the above STR-1 primers, STR-2 primers, STR-3 primers, STR-4 primers, STR-5 primers, and STR-6 primers are all 10 μM, and the primer sequences are shown in the following table:

The PCR amplification reaction solution is a mixture of the following reagents: TaqDNA polymerase (5U/μL), Tris-HCl (100mM, pH 8.8 at 25°C), KCl (500mM), ethylphenyl polyethylene glycol (0.8 % (v/v)), _MgCl2 (25 mM), dNTPs (10 mM), deionized water.

The PCR amplification reaction solution was stored at -20°C; the LIZ-500 molecular weight internal standard was stored at -20°C; and the deionized formamide was stored at 2-8°C.

The above-mentioned kit also includes instructions for use.

Example 3 Using the system of Example 1 and the kit of Example 2 to predict the risk of the subject suffering from laryngeal squamous cell carcinoma

Subject: Male, 59 years old, visiting the Department of Otolaryngology, China-Japan Friendship Hospital of Jilin University, fully informed of the purpose and use of the examination, signed an informed consent form, and collected 1mL of anticoagulant through peripheral veins .

Using the kit of Example 2, the following steps were performed according to the method described in the kit instructions:

(1) Extract DNA from the sample: extract genomic DNA from blood using a DNA extraction kit;

(2) PCR reaction

(2-1) Take out the STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, STR-6 primer, and PCR amplification reaction solution from the refrigerator, and equilibrate to room temperature. After the components are fully dissolved, centrifuge quickly for 10 seconds;

(2-2) Take 100ng of sample DNA, add 60μL of PCR amplification reaction solution, add deionized water to make up to 115.2μL, mix well, centrifuge quickly for 10 seconds, and then divide the mixture into 6 cells at 19.2μL/well PCR reaction tube;

(2-3) Add STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, and STR-6 primer to step (2-2) at 0.8 μL/well respectively. 6 PCR reaction tubes; cover the PCR reaction tube caps, record the sample loading situation, centrifuge quickly for 10 seconds, then transfer the PCR reaction tubes to the corresponding position of the sample tank of the PCR amplifier, record the placement sequence, and start PCR amplification Amplification reaction conditions: 95°C for 3 minutes; 95°C for 30 seconds, 60°C for 30 seconds, 72°C for 30 seconds, 10 cycles; 95°C for 30 seconds, 55°C for 30 seconds, 72°C for 30 seconds, 20 cycles ; 6 minutes at 72°C to obtain 6 sets of PCR amplification products;

(3) STR fragment analysis

(3-1) Take 990 μL of deionized formamide, add 10 μL of LIZ-500 molecular weight internal standard, mix well, centrifuge quickly for 10 seconds, add 10 μL/well to sequencing reaction tubes, and centrifuge quickly for 10 seconds;

(3-2) Add 1 μL/well of the above-mentioned 6 sets of PCR amplification products to 6 sequencing reaction tubes, and centrifuge quickly for 10 seconds; then transfer the sequencing reaction tubes to the corresponding position of the sample tank of the PCR amplifier, 98°C Heating for 5 minutes. Immediately after the end of the program, the sequencing reaction tube was placed on the ice-water mixture and rapidly cooled to 0 °C and centrifuged rapidly for 10 seconds; then the sequencing reaction tube was transferred to the corresponding position of the sample tank of the STR locus fragment analyzer, and recorded and placed sequence, perform fragment analysis and detection;

(4) Analysis and judgment of results

(4-1) According to the fragment analysis results, record the fragment lengths of the two alleles of STR-1, STR-2, STR-3, STR-4, STR-5, and STR-6 respectively: STR-1 two The smaller fragment length value of the two alleles is recorded as L ₁ , the larger fragment length value of the two alleles of STR-1 is recorded as L ₂ ; the smaller fragment length of the two alleles of STR-2 is recorded as L 2 ; The value is recorded as L ₃ , the larger fragment length value of the two alleles of STR-2 is recorded as L ₄ ; the smaller fragment length value of the two alleles of STR-3 is recorded as L ₅ , and the two STR-3 alleles are recorded as L 5 . The larger fragment length value of the two alleles is recorded as L ₆ ; the smaller fragment length value of the two alleles of STR-4 is recorded as L ₇ , and the larger fragment length of the two alleles of STR-4 is recorded as L 7 . The value is recorded as L ₈ ; the smaller fragment length value of the two alleles of STR-5 is recorded as L ₉ , and the larger fragment length value of the two alleles of STR-5 is recorded as L ₁₀ ; The smaller fragment length value of the two alleles was recorded as L ₁₁ , and the larger fragment length value of the two alleles of STR-6 was recorded as L ₁₂ ; the results showed: L ₁ =260.25, L ₂ =260.25, L ₃ =402.1, L4=402.1, L5= _229.03 , L6= _246.15 , _L7 = _386.85 , L8= _400.5 , L9= _312.16 , L10= _318.59 , _L11 =256.99, L12= _260.51 .

(4-2) Calculated according to the length of the fragment and the following formula, denoted as X ₁ -X ₁₂ , where round represents rounding to an integer:

X ₁ =round[(L ₁ -191)/3]=23; X ₂ =round[(L ₂ -191)/3]=23;

X ₃ =round(L ₃ -379)=23; X ₄ =round(L ₄ -379)=23;

X ₅ =round[(L ₅ -202)/2]=14; X ₆ =round[(L ₆ -202)/2]=22;

X ₇ =round[(L ₇ -359)/2]=14; X ₈ =round[(L ₈ -359)/2]=21;

X ₉ =round[(L ₉ -278)/2]=17; X ₁₀ =round[(L ₁₀ -278)/2]=20;

X ₁₁ =round[(L ₁₁ -200)/4]=14; X ₁₂ =round[(L ₁₂ -200)/4]=15;

The patient was male, X ₁₃ =1.

(4-3) Using the computer running the laryngeal squamous cell carcinoma susceptibility prediction system described in Example 1, predict the susceptibility of the subject to suffer from laryngeal squamous cell carcinoma:

The age, gender, and repetition times of the short tandem sequence of the STR site are input into the system through the data input module, and the result of the discriminant function is calculated through the data calculation module:

First discriminant function F _LC =10.750X ₁ -8.272X ₂ +14.088X ₃ +106.284X ₄ -0.418X ₅ +2.056X ₆ +2.288X ₇ +1.131X ₈ -0.466X ₉ -4.985X ₁₀ +2.407X ₁₁ +7.072X ₁₂ -15.282X ₁₃ -1550.869＝1386.718

Second discriminant function F _LN =11.048X ₁ -8.564X ₂ +14.325X ₃ +108.309X ₄ -0.776X ₅ +1.944X ₆ +2.347X ₇ +0.795X ₈ -0.807X ₉ -4.929X ₁₀ +2.523X ₁₁ +6.583X ₁₂ -18.780X ₁₃ -1581.097＝1381.062

After the analysis, discrimination and result output module compares the F _LC value and the F _LN value, F _LC > F _LN , and outputs the prediction result of "the probability of the subject suffering from laryngeal squamous cell carcinoma ≥ 82.4%".

The subject underwent a biopsy of the laryngeal tumor after seeing a doctor, and the pathological examination was confirmed to be laryngeal squamous cell carcinoma, and the clinical diagnosis result was consistent with the prediction result of the kit of the present invention.

Example 4 Using the system of Example 1 and the kit of Example 2 to predict the risk of the subject suffering from laryngeal squamous cell carcinoma

Subject: female, 72 years old, visiting the Department of Otolaryngology, China-Japan Friendship Hospital of Jilin University, fully informed of the purpose and use of the examination, signed an informed consent form, and collected 1mL of anticoagulant through peripheral veins .

Referring to the prediction method of Example 3, the same processing and testing were performed on the blood samples, and the results showed: L ₁ =268.17, L ₂ =268.17, L ₃ =404.17, L ₄ =404.17, L ₅ =228.76, L ₆ =228.76, L ₇ =386.85, L ₈ =390.67, L ₉ =311.75, L ₁₀ =311.75, L ₁₁ =260.20, L ₁₂ =263.85.

Calculated according to the fragment length and the following formula, denoted as X ₁ -X ₁₂ , where round represents rounding to an integer:

X ₁ =round[(L ₁ -191)/3]=26; X ₂ =round[(L ₂ -191)/3]=26;

X ₃ =round(L ₃ -379)=25; X ₄ =round(L ₄ -379)=25;

X ₅ =round[(L ₅ -202)/2]=13; X ₆ =round[(L ₆ -202)/2]=13;

X ₇ =round[(L ₇ -359)/2]=14; X ₈ =round[(L ₈ -359)/2]=16;

X ₉ =round[(L ₉ -278)/2]=17; X ₁₀ =round[(L ₁₀ -278)/2]=17;

X ₁₁ =round[(L ₁₁ -200)/4]=15; X ₁₂ =round[(L ₁₂ -200)/4]=16;

The patient was female, X ₁₃ =0.

Use the computer running the laryngeal squamous cell carcinoma susceptibility prediction system described in Example 1 to predict the susceptibility of the subject to suffer from laryngeal squamous cell carcinoma:

The age, gender, and repetition times of the short tandem sequence of the STR site are input into the system through the data input module, and the result of the discriminant function is calculated through the data calculation module:

First discriminant function F _LC =10.750X ₁ -8.272X ₂ +14.088X ₃ +106.284X ₄ -0.418X ₅ +2.056X ₆ +2.288X ₇ +1.131X ₈ -0.466X ₉ -4.985X ₁₀ +2.407X ₁₁ +7.072X ₁₂ -15.282X ₁₃ -1550.869＝1650.871

Second discriminant function F _LN =11.048X ₁ -8.564X ₂ +14.325X ₃ +108.309X ₄ -0.776X ₅ +1.944X ₆ +2.347X ₇ +0.795X ₈ -0.807X ₉ -4.929X ₁₀ +2.523X ₁₁ +6.583X ₁₂ -18.780X ₁₃ -1581.097＝1655.76

The analysis, discrimination and result output module compares the F _LC value and the F _LN value, F _LC ≤ F _LN , and outputs the prediction result of "the probability of the subject not suffering from laryngeal squamous cell carcinoma ≥ 83.3%".

The subject was diagnosed with vocal cord polyps after seeing a doctor, and the clinical diagnosis results were consistent with the predicted results of the kit of the present invention.

The above are the preferred embodiments of the present invention, and are not intended to limit the present invention. It should be pointed out that any changes, modifications, substitutions and alterations (such as: increase, decrease, change, etc.) made without departing from the principle and purpose of the present invention STR sites, using cells or tissues from other sources of the subject, using other similar statistical methods, etc.) should all be included within the scope of protection of the present invention.

SEQUENCE LISTING

<110> Jilin University

<120> A kit and system for predicting susceptibility to laryngeal squamous cell carcinoma

<130> DI18-8168-XC47

<160> 12

<170> PatentIn version 3.3

<210> 1

<211> 18

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(18)

<223> STR-1 primer upstream primer

<400> 1

agggctggga agggtcta 18

<210> 2

<211> 19

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(19)

<223> STR-1 primer downstream primer

<400> 2

ggagaaccat cctcaccct 19

<210> 3

<211> 20

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(20)

<223> STR-2 primer upstream primer

<400> 3

cgcctccaag aatgtaagtg 20

<210> 4

<211> 22

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(22)

<223> STR-2 primer downstream primer

<400> 4

aactcaagtc tatgcttcac cc 22

<210> 5

<211> 21

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(21)

<223> STR-3 primer upstream primer

<400> 5

ggtttccatt gtagcatctt g 21

<210> 6

<211> 20

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(20)

<223> STR-3 primer downstream primer

<400> 6

gcctggttgt ttccgtagta 20

<210> 7

<211> 20

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(20)

<223> STR-4 primer upstream primer

<400> 7

tctgttgggt gtttgggata 20

<210> 8

<211> 20

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(20)

<223> STR-4 primer downstream primer

<400> 8

ttacattgtc ggtctggtcc 20

<210> 9

<211> 20

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(20)

<223> STR-5 primer upstream primer

<400> 9

atctcagtct ccccaagtgc 20

<210> 10

<211> 20

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(20)

<223> STR-5 primer downstream primer

<400> 10

tccttcaaga taaccaccga 20

<210> 11

<211> 22

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(22)

<223> STR-6 primer upstream primer

<400> 11

tcggttgtag gtattatcac gg 22

<210> 12

<211> 20

<212> DNA

<213> Artificial

<220>

<221> primer_bind

<222> (1)..(20)

<223> STR-6 primer downstream primer

<400> 12

tgcccccatag gttttgaact 20

Claims (7)

1. A kit for predicting susceptibility to laryngeal squamous cell carcinoma, comprising the following components: STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, STR-6 primer , the STR-1 primers, STR-2 primers, STR-3 primers, STR-4 primers, STR-5 primers, and STR-6 primers are respectively used to amplify the target fragments containing the following short tandem sequences to determine the following short The number of repetitions of the tandem sequence: Site code starting point gene short tandem sequence STR-1 X chromosome, position 66657655 AR CAG STR-2 Chromosome 4, position 55633758 Bat-25 T STR-3 Chromosome 5, position 111646983 D5S346 GT STR-4 Chromosome 6, position 151806531 ER1 TA STR-5 Chromosome 14, position 64253561 ER2 TG STR-6 Chromosome 4, position 154587748 FGA AAAG The sequences of the STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer and STR-6 primer are as follows:

Wherein, the kit further includes: PCR amplification reaction solution, LIZ-500 molecular weight internal standard, deionized formamide, and instructions for use, The instructions for use describe a method of using the kit for predicting susceptibility to laryngeal squamous cell carcinoma, which includes the following steps: (1) Extract sample DNA; (2) PCR reaction (2-1) Take out the STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, STR-6 primer, and PCR amplification reaction solution from the refrigerator, and equilibrate to room temperature. After the components are fully dissolved, centrifuge quickly for 10 seconds; (2-2) Take 30-300ng of sample DNA, add 60μL of PCR amplification reaction solution, add deionized water to make up to 115.2μL, mix well, centrifuge quickly for 10 seconds, and then dispense the mixture at 19.2μL/well to 6 PCR reaction tubes; (2-3) Add STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, and STR-6 primer to step (2-2) at 0.8 μL/well respectively. 6 PCR reaction tubes; cover the PCR reaction tube caps, record the sample loading situation, centrifuge quickly for 10 seconds, then transfer the PCR reaction tubes to the corresponding position of the sample tank of the PCR amplifier, record the placement sequence, and start PCR amplification Amplification reaction conditions: 95°C for 3 minutes; 95°C for 30 seconds, 60°C for 30 seconds, 72°C for 30 seconds, 10 cycles; 95°C for 30 seconds, 55°C for 30 seconds, 72°C for 30 seconds, 20 cycles ; 6 minutes at 72°C to obtain 6 sets of PCR amplification products; (3) STR fragment analysis (3-1) Take 990 μL of deionized formamide, add 10 μL of LIZ-500 molecular weight internal standard, mix well, centrifuge quickly for 10 seconds, add 10 μL/well to sequencing reaction tubes, and centrifuge quickly for 10 seconds; (3-2) Add 1 μL/well of the above-mentioned 6 sets of PCR amplification products to 6 sequencing reaction tubes, and centrifuge quickly for 10 seconds; then transfer the sequencing reaction tubes to the corresponding position of the sample tank of the PCR amplifier, 98°C Heating for 5 minutes. Immediately after the end of the program, the sequencing reaction tube was placed on the ice-water mixture and rapidly cooled to 0 °C and centrifuged rapidly for 10 seconds; then the sequencing reaction tube was transferred to the corresponding position of the sample tank of the STR locus fragment analyzer, and recorded and placed sequence, perform fragment analysis and detection; (4) Analysis and judgment of results (4-1) According to the fragment analysis results, record the fragment lengths of the two alleles of STR-1, STR-2, STR-3, STR-4, STR-5, and STR-6 respectively: The smaller fragment length value of the two alleles of STR-1 is recorded as L ₁ , and the larger fragment length value of the two alleles of STR-1 is recorded as L ₂ ; The smaller fragment length value of the two alleles of STR-2 is recorded as L ₃ , and the larger fragment length value of the two alleles of STR-2 is recorded as L ₄ ; The smaller fragment length value of the two alleles of STR-3 is recorded as L ₅ , and the larger fragment length value of the two alleles of STR-3 is recorded as L ₆ ; The smaller fragment length value of the two alleles of STR-4 is recorded as L ₇ , and the larger fragment length value of the two alleles of STR-4 is recorded as L ₈ ; The smaller fragment length value of the two alleles of STR-5 is recorded as L ₉ , and the larger fragment length value of the two alleles of STR-5 is recorded as L ₁₀ ; The smaller fragment length value of the two alleles of STR-6 is recorded as L ₁₁ , and the larger fragment length value of the two alleles of STR-6 is recorded as L ₁₂ ; (4-2) The number of repetitions of the short tandem sequence is calculated according to the fragment length and the following formula, and is denoted as X ₁ -X ₁₂ , where round represents rounding to an integer: X ₁ =round[(L ₁ -191)/3]; X ₂ =round[(L ₂ -191)/3]; X ₃ =round(L ₃ -379); X ₄ =round(L ₄ -379); X ₅ =round[(L ₅ -202)/2]; X ₆ =round[(L ₆ -202)/2]; X ₇ =round[(L ₇ -359)/2]; X ₈ =round[(L ₈ -359)/2]; X ₉ =round[(L ₉ -278)/2]; X ₁₀ =round[(L ₁₀ -278)/2]; X ₁₁ =round[(L ₁₁ -200)/4]; X ₁₂ =round[(L ₁₂ -200)/4]; (4-3) Substitute the repetition times of the above short series sequence into the preset discriminant function: F _LC = 10.750X ₁ -8.272X ₂ +14.088X ₃ +106.284X ₄ -0.418X ₅ +2.056X ₆ +2.288X ₇ +1.131X ₈ -0.466X ₉ -4.985X ₁₀ +2.407X ₁₁ +7.072X ₁₂ -15.282X ₁₃ -1550.869 F _LN = 11.048X ₁ -8.564X ₂ +14.325X ₃ +108.309X ₄ -0.776X ₅ +1.944X ₆ +2.347X ₇ +0.795X ₈ -0.807X ₉ -4.929X ₁₀ +2.523X ₁₁ +6.583X ₁₂ -18.780X ₁₃ -1581.097 Wherein, if the subject is female, X ₁₃ =0; if the subject is male, X ₁₃ =1; (4-4) Prediction of susceptibility to laryngeal squamous cell carcinoma: Comparing the F _LC value and the F _LN value, if F _LC > F _LN , the probability of the subject suffering from laryngeal squamous cell carcinoma is predicted to be ≥82.4%; if F _LC ≤ F _LN , it is predicted that the subject will not suffer from laryngeal squamous cell carcinoma The chance of cell carcinoma is ≥83.3%. 2. The kit for predicting susceptibility to laryngeal squamous cell carcinoma according to claim 1, wherein the STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer and STR-6 primers were used at a concentration of 10 μM. 3. laryngeal squamous cell carcinoma susceptibility prediction kit as claimed in claim 1, is characterized in that: described PCR amplification reaction solution is the mixed solution of following reagent: TaqDNA polymerase 5U/μL, Tris-HCl 100mM, KCl 500 mM, ethylphenyl polyethylene glycol 0.8 vol%, MgCl ₂ 25 mM, dNTP 10 mM, deionized water; the pH of Tris-HCl is 8.8 at 25°C. 4 . The kit for predicting susceptibility to laryngeal squamous cell carcinoma according to claim 1 , wherein the sample is whole blood, oral swab or oral tissue of the subject. 5 . 5. The application of the laryngeal squamous cell carcinoma susceptibility prediction kit according to any one of claims 1 to 4 in the preparation of a laryngeal squamous cell carcinoma diagnostic product. 6. A laryngeal squamous cell carcinoma susceptibility prediction system, characterized in that it comprises: A device for obtaining the number of repetitions of short tandem sequences of the following STR loci in sample DNA: Site code starting point gene short tandem sequence STR-1 X chromosome, position 66657655 AR CAG STR-2 Chromosome 4, position 55633758 Bat-25 T STR-3 Chromosome 5, position 111646983 D5S346 GT STR-4 Chromosome 6, position 151806531 ER1 TA STR-5 Chromosome 14, position 64253561 ER2 TG STR-6 Chromosome 4, position 154587748 FGA AAAG ; A data processing and determination device, which includes the following modules: The data input module is used to input the age, gender, and repetition times of the short tandem sequence of the STR locus; The database management module is used for the operation management of data storage, modification, deletion, query and printing; The data calculation module is used to calculate the discriminant function result according to the repetition times of the short tandem sequence of the STR site in the data input module; The analysis and discrimination and result output module is used to compare the discriminant function results, so as to predict the susceptibility of laryngeal squamous cell carcinoma, and output the results, Wherein, the number of repetitions X ₁ -X ₁₂ of the short tandem sequence of the STR site of the sample DNA is obtained by using the kit according to any one of claims 1 to 5; and in: The discriminant function includes: First discriminant function F _LC =10.750X ₁ -8.272X ₂ +14.088X ₃ +106.284X ₄ -0.418X ₅ +2.056X ₆ +2.288X ₇ +1.131X ₈ -0.466X ₉ -4.985X ₁₀ +2.407X ₁₁ +7.072X ₁₂ -15.282X ₁₃ -1550.869 Second discriminant function F _LN =11.048X ₁ -8.564X ₂ +14.325X ₃ +108.309X ₄ -0.776X ₅ +1.944X ₆ +2.347X ₇ +0.795X ₈ -0.807X ₉ -4.929X ₁₀ +2.523X ₁₁ +6.583X ₁₂ -18.780X ₁₃ -1581.097 In the discriminant function, X ₁ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-1; X ₂ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-1; X ₃ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-2; X ₄ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-2; X ₅ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-3; X ₆ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-3; X ₇ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-4; X ₈ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-4; X ₉ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-5; X ₁₀ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-5; X ₁₁ is the number of repetitions of the short tandem sequence of the smaller fragment in the two alleles of STR-6; X ₁₂ is the number of repetitions of the short tandem sequence of the larger fragment in the two alleles of STR-6; If the subject is female, X ₁₃ =0; if the subject is male, X ₁₃ =1; Among them, X ₁ ^- X ₁₂ are calculated according to the length of the segment and as follows, where round represents rounding to an integer: X ₁ =round[(L ₁ -191)/3]; X ₂ =round[(L ₂ -191)/3]; X ₃ =round(L ₃ -379); X ₄ =round(L ₄ -379); X ₅ =round[(L ₅ -202)/2]; X ₆ =round[(L ₆ -202)/2]; X ₇ =round[(L ₇ -359)/2]; X ₈ =round[(L ₈ -359)/2]; X ₉ =round[(L ₉ -278)/2]; X ₁₀ =round[(L ₁₀ -278)/2]; X ₁₁ =round[(L ₁₁ -200)/4]; X ₁₂ =round[(L ₁₂ -200)/4]; In X ₁ ^- X ₁₂ , L ₁ is the smaller fragment length value of the two alleles of STR-1, and L ₂ is the larger fragment length value of the two alleles of STR-1; L ₃ is the smaller fragment length value of the two alleles of STR-2, and L ₄ is the larger fragment length value of the two alleles of STR-2; L ₅ is the smaller fragment length value of the two alleles of STR-3, and L ₆ is the larger fragment length value of the two alleles of STR-3; L ₇ is the smaller fragment length value of the two alleles of STR-4, and L ₈ is the larger fragment length value of the two alleles of STR-4; L ₉ is the smaller fragment length value of the two alleles of STR-5, and L ₁₀ is the larger fragment length value of the two alleles of STR-5; L ₁₁ is the smaller fragment length value of the two alleles of STR-6, and L ₁₂ is the larger fragment length value of the two alleles of STR-6; The analysis, discrimination and result output module compares the calculation result of the first discriminant function F _LC with the calculation result of the second discriminant function F _LN , and if F _LC > F _LN , it outputs “The subject suffers from laryngeal squamous cells. If F _LC ≤ F _LN , output the prediction result of “the probability that the subject does not suffer from laryngeal squamous cell carcinoma ≥ 83.3%”. 7. The application of the laryngeal squamous cell carcinoma susceptibility prediction system of claim 6 in the preparation of laryngeal squamous cell carcinoma prediction products or laryngeal squamous cell carcinoma diagnostic products.

Images