CN108676890B - Female breast malignant tumor susceptibility prediction kit and system - Google Patents
Female breast malignant tumor susceptibility prediction kit and system Download PDFInfo
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Abstract
The invention relates to a female breast malignant tumor susceptibility prediction kit, which comprises the following components: STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer and STR-6 primer; further, it may further include: PCR amplification reaction liquid, LIZ-500 molecular weight internal standard and deionized formamide. The kit for predicting the susceptibility of the female breast malignant tumor can be used for diagnosing the female breast malignant tumor and predicting the susceptibility. The invention also provides a system for predicting the susceptibility of the female breast malignant tumor.
Description
Technical Field
The present invention relates to the field of biomedicine. In particular to a female breast malignant tumor susceptibility prediction kit and a female breast malignant tumor susceptibility prediction system. More specifically, the invention relates to a kit for detecting STR of female breast malignant tumor susceptibility related genes by Short Tandem Repeat (STR) locus fragment analysis method, and early warning is carried out on female breast malignant tumor susceptibility of a detected object by combining with discriminant analysis statistical method.
Background
The tumor is a disease closely related to genetic genes, the molecular genetics basis of the tumor is researched, and further a tumor specific genetics marker is provided, so that the tumor specific genetics marker is expected to provide a simple and feasible method for common detection, clinical diagnosis, personalized treatment, disease tracking after recovery and the like. However, the individual differences of patients, the intercrossing of related biomolecular events at different stages of development, etc. all bring great difficulties to the work.
The female breast malignant tumor is mainly breast cancer from a breast duct and lobular epithelium, and accounts for more than 95 percent of the female breast malignant tumor; secondly, soft tissue sarcoma from mammary mesenchymal tissue, etc., less than 5%. The occurrence of this type of disease is related to endocrine factors, diet and obesity, radiation exposure, milk factor, etc. Therefore, the method can predict the susceptibility of the breast malignant tumor of the human population to be detected, is favorable for improving the risk awareness of the disease, reduces the pathogenesis by adjusting the diet, the working environment and the like, and is also favorable for the early discovery and early treatment of the patient.
A large number of studies have shown that genetic polymorphisms of tumor-associated genes play a key role in the development of malignant tumors. However, the development of tumors is a very complex process, and the diagnosis of the disease using changes in a single molecular genetic marker is clearly impossible and not scientific. In the prior art, accurate early warning on tumor susceptibility cannot be performed only through genetic information, and the current early identification and prediction method for tumors needs to be improved. The invention relates to a kit for early warning susceptibility of female breast malignant tumors by jointly detecting a plurality of STR loci with high relevance to the occurrence of female breast malignant tumors through an STR locus fragment analysis method and combining a discriminant analysis statistical method.
Disclosure of Invention
In order to solve the problems in the prior art, the invention relates to a method for jointly detecting a plurality of STR loci with high relevance to the occurrence of female breast malignant tumors by an STR locus fragment analysis method, and early warning is carried out on the susceptibility of the female breast malignant tumors by combining a discriminant analysis statistical method.
The present invention has been completed based on the following findings of the inventors: the inventor discovers that the repetition times of short tandem sequences of each independent STR locus has no significant correlation with the female breast malignant tumor of the detected object, and the combination of the repetition times of the short tandem sequences of certain specific STR loci has close relation with the female breast malignant tumor of the detected object by analyzing STRs of the female breast malignant tumor detected objects and the genome DNA of healthy control detected objects and verifying the STRs in a large number of female breast malignant tumor samples and control samples.
To this end, the present invention proposes a set of isolated STR sites that have a high association with the development of female breast malignancies. According to an embodiment of the present invention, these isolated STR loci comprise the nucleotide sequences shown as STR-1 to STR-6 (Table 1). By using the separated STR loci as reference, the susceptibility of female breast malignant tumor can be effectively predicted.
TABLE 1
| Locus code | Starting position | Belonging 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 above detailed description of STR sites, those skilled in the art can log in relevant databases (such as GeneBank, Nucleotide, etc.) to obtain the details, which are not described herein. The inventor surprisingly finds that the short tandem sequence repeat times of each STR locus are obtained by analyzing the cell genome of a detected object, and a statistical analysis method such as discriminant analysis and the like is carried out by taking the repeat times as independent variables, so that early warning can be carried out on the female breast malignant tumor susceptibility.
On the basis, one of the technical problems solved by the invention is to provide a kit for predicting the susceptibility of female breast malignant tumors, which comprises the following components: STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer and STR-6 primer, wherein the primers are respectively used for amplifying target fragments containing short tandem sequences listed in Table 1 so as to determine the repetition times of the short tandem sequences.
Preferably, the kit for predicting susceptibility to breast cancer of a female of the present invention further comprises: PCR amplification reaction liquid, LIZ-500 molecular weight internal standard and deionized formamide.
In the kit for predicting susceptibility to female breast cancer of the present invention, preferably, 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 shown in table 2 below, and more preferably, the concentrations of the primers are all 10 μ M:
TABLE 2
In table 2, HEX, FAM, and ROX are all fluorophores labeling the 5' end, HEX is hexachloro-6-methylfluorescein, FAM is 6-carboxyfluorescein, and ROX is ROX reference dye.
In the kit for predicting susceptibility to female breast malignant tumor of the present invention, preferably, the PCR amplification reaction solution is a mixed solution of the following reagents: TaqDNA polymerase (5U/. mu.L), Tris-HCl (100mM, pH 8.8 at 25 ℃), KCl (500mM), ethylphenylpolyethyleneglycol (0.8% (v/v)), MgCl2(25mM), dNTP (10mM), deionized water.
More preferably, the PCR amplification reaction solution is stored at-20 ℃.
In the kit for predicting susceptibility to female breast malignant tumor of the present invention, preferably, the LIZ-500 molecular weight internal standard can be preserved at-20 ℃;
in the kit for predicting susceptibility to female breast malignant tumor of the present invention, preferably, the deionized formamide can be stored at 2-8 ℃.
Preferably, the kit for predicting susceptibility to breast cancer in women of the present invention further comprises instructions for use.
The application instruction describes a use method of the kit for predicting susceptibility to female breast malignant tumor, which comprises the following steps:
(1) extracting sample DNA;
(2) PCR reaction
(2-1) taking out the STR-1 primer, the STR-2 primer, the STR-3 primer, the STR-4 primer, the STR-5 primer, the STR-6 primer and the PCR amplification reaction solution from a refrigerator, balancing to room temperature, fully dissolving each component, and respectively and rapidly centrifuging for 10 seconds;
(2-2) adding 30-300ng of sample DNA into 60 mu L of PCR amplification reaction solution, adding deionized water to supplement to 115.2 mu L, fully and uniformly mixing, quickly centrifuging for 10 seconds, and subpackaging the mixed solution into 6 PCR reaction tubes according to 19.2 mu L/hole;
(2-3) respectively adding an STR-1 primer, an STR-2 primer, an STR-3 primer, an STR-4 primer, an STR-5 primer and an STR-6 primer into the 6 PCR reaction tubes in the step (2-2) according to 0.8 mu L/hole; covering a PCR reaction tube cover, recording the sample adding condition, quickly centrifuging for 10 seconds, then transferring the PCR reaction tube to a corresponding position of a sample groove of a PCR amplification instrument, recording the placing sequence, and starting the PCR amplification reaction; the amplification reaction conditions are as follows: 3 minutes at 95 ℃; 30 seconds at 95 ℃, 30 seconds at 60 ℃ and 30 seconds at 72 ℃ for 10 cycles; 30 seconds at 95 ℃, 30 seconds at 55 ℃, 30 seconds at 72 ℃ and 20 cycles; 6 groups of PCR amplification products are obtained at 72 ℃ for 6 minutes;
(3) STR fragment analysis
(3-1) adding 990 mu L of deionized formamide into 10 mu L of LIZ-500 molecular weight internal standard, fully and uniformly mixing, quickly centrifuging for 10 seconds, respectively adding into a sequencing reaction tube according to 10 mu L/hole, and quickly centrifuging for 10 seconds;
(3-2) adding the 6 groups of PCR amplification products into 6 sequencing reaction tubes according to 1 mu L/hole respectively, and quickly centrifuging for 10 seconds; then transferring the sequencing reaction tube to a corresponding position of a sample tank of a PCR (polymerase chain reaction) amplification instrument, heating at 98 ℃ for 5 minutes, immediately placing the sequencing reaction tube on an ice-water mixture after the program is finished, rapidly cooling to 0 ℃, and rapidly centrifuging for 10 seconds; then transferring the sequencing reaction tube to a corresponding position of a sample groove of an STR locus fragment analyzer, recording the placement sequence, and performing fragment analysis detection;
(4) analysis and determination of results
(4-1) respectively recording the fragment lengths of two alleles at each site of STR-1, STR-2, STR-3, STR-4, STR-5 and STR-6 according to the fragment analysis result:
the length of the smaller of the two STR-1 alleles is recorded as L1And the length of the larger fragment of the two STR-1 alleles is designated as L2；
The length of the smaller fragment of the two STR-2 alleles is recorded as L3，STR-The larger of the two alleles has a length L4；
The length of the smaller fragment of the two STR-3 alleles is recorded as L5And the length of the larger fragment of the two STR-3 alleles is marked as L6；
The length of the smaller fragment of the two STR-4 alleles is recorded as L7And the length of the larger fragment of the two STR-4 alleles is marked as L8；
The length of the smaller of the two STR-5 alleles is recorded as L9And the length of the larger fragment of the two STR-5 alleles is designated as L10；
The length of the smaller fragment of the two STR-6 alleles was designated L11And the length of the larger fragment of the two STR-6 alleles is marked as L12；
(4-2) the number of repetitions of the short tandem sequence is calculated from the fragment length and the following formula, and is denoted as X1-X12Where round stands for rounded integer:
X1＝round[(L1-191)/3]；X2＝round[(L2-191)/3]；
X3＝round(L3-379)；X4＝round(L4-379)；
X5＝round[(L5-202)/2]；X6＝round[(L6-202)/2]；
X7＝round[(L7-359)/2]；X8＝round[(L8-359)/2]；
X9＝round[(L9-278)/2]；X10＝round[(L10-278)/2]；
X11＝round[(L11-200)/4]；X12＝round[(L12-200)/4]；
(4-3) substituting the number of the short tandem sequence repetitions into a preset discriminant function:
FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160
FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413
(4-4) prediction of susceptibility to female breast malignancy:
comparison FBCValue sum FBNValue if FBC>FBNThen the probability of the female breast malignant tumor of the detected object is predicted to be more than or equal to 90.0 percent; if FBC≤FBNAnd predicting that the probability that the detected object does not suffer from the female breast malignant tumor is more than or equal to 77.8 percent.
In the present invention,
preferably, the sample DNA extracted in step (1) can be performed using a commercially available genomic DNA extraction kit according to the kit instructions. The sample may be whole blood of a subject.
Preferably, the rotational speed of all the centrifuges in the method of use is preferably 3000 g/min.
The probability of suffering from female breast malignant tumor in the invention is the sum of the probability of suffering from female breast malignant tumor already and the probability of suffering from female breast malignant tumor in the future. Therefore, the method can be used for diagnosing the female breast malignant tumor; the risk early warning method can also be used for risk early warning of the future female breast malignant tumor, can assist a detected object to carry out risk prevention, and reduces the disease probability of the female breast malignant tumor through the modes of medicine conditioning, change of life and rest, diet rule, regular physical examination and the like.
The second technical problem solved by the invention is to provide a method for predicting the susceptibility of female breast malignant tumor, namely, the kit is used and the method is operated according to the instruction.
The invention solves the third technical problem by providing the application of the female breast malignant tumor susceptibility prediction kit in preparing female breast malignant tumor diagnosis products.
The fourth technical problem to be solved by the present invention is to provide a system for predicting susceptibility to female breast malignant tumor, which comprises:
a device for obtaining the repeat times of the STR locus short tandem sequence of the sample DNA;
data processing and decision device, comprising the following modules:
the data input module is used for inputting the age, the sex and the STR locus short tandem sequence repetition times of the detected object;
the database management module is used for the operation management of data storage, modification, deletion, inquiry and printing;
the data calculation module is used for calculating a discrimination function result according to the repeat times of the STR locus short serial sequence in the data input module;
and the analysis, discrimination and result output module is used for comparing the discrimination function results so as to predict the susceptibility of the female breast malignant tumor and output the result.
Wherein the content of the first and second substances,
the number of times of the STR locus short tandem sequence repetition is 6 pairs of the number of times of the STR locus short tandem sequence repetition:
| locus code | Starting position | Belonging 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 FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160
Second discrimination function FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413
In the case of the discriminant function,
X1the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-1;
X2the number of repeats of the short tandem sequence for the larger of the two alleles of STR-1;
X3the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-2;
X4the number of repeats of the short tandem sequence for the larger of the two alleles of STR-2;
X5the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-3;
X6the number of repeats of the short tandem sequence for the larger of the two alleles of STR-3;
X7the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-4;
X8the number of repeats of the short tandem sequence for the larger of the two alleles of STR-4;
X9the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-5;
X10the number of repeats of the short tandem sequence for the larger of the two alleles of STR-5;
X11the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-6;
X12the number of repeats of the short tandem sequence for the larger of the two alleles of STR-6;
wherein, X1 -X12Calculated from the segment length and the formulaMedium round stands for rounded integers:
X1＝round[(L1-191)/3]；X2＝round[(L2-191)/3]；
X3＝round(L3-379)；X4＝round(L4-379)；
X5＝round[(L5-202)/2]；X6＝round[(L6-202)/2]；
X7＝round[(L7-359)/2]；X8＝round[(L8-359)/2]；
X9＝round[(L9-278)/2]；X10＝round[(L10-278)/2]；
X11＝round[(L11-200)/4]；X12＝round[(L12-200)/4]；
X1 -X12in, L1Is the smaller segment length value, L, of the two alleles of STR-12Is the larger fragment length value of the two alleles of STR-1;
L3is the smaller segment length value, L, of the two alleles of STR-24Is the larger fragment length value of the two alleles of STR-2;
L5is the smaller segment length value, L, of the two alleles of STR-36Is the larger fragment length value of the two alleles of STR-3;
L7is the smaller segment length value, L, of the two alleles of STR-48Is the larger fragment length value of the two alleles of STR-4;
L9is the smaller segment length value, L, of the two alleles of STR-510Is the larger fragment length value of the two alleles of STR-5;
L11is the smaller segment length value, L, of the two STR-6 alleles12Is the larger fragment length value of the two alleles of STR-6;
the analysis discrimination and result output module outputs a first discrimination function FBCAnd the second judgmentOther function FBNIf F is the result of the calculation ofBC>FBNOutputting a prediction result that the probability of the female breast malignant tumor of the detected object is more than or equal to 90.0%; if FBC≤FBNAnd outputting a prediction result that the probability that the detected object does not suffer from the female breast malignant tumor is more than or equal to 77.8 percent.
The device for obtaining the repetition times of the STR locus short tandem sequence of the sample DNA can comprise an STR locus fragment analyzer, a PCR amplification instrument and the like; the data processing and determining device may be a computer or the like.
The fifth technical problem to be solved by the invention is to provide the application of the female breast malignant tumor susceptibility prediction system in the preparation of female breast malignant tumor prediction products, female breast malignant tumor diagnosis products and breast health auxiliary products.
The sixth technical problem to be solved by the invention is to provide a female breast malignant tumor prediction product, a female breast malignant tumor diagnosis product or a breast health auxiliary product, which comprises the female breast malignant tumor susceptibility prediction system.
The test material used in the present invention is human genomic DNA, which theoretically does not change during the life of a human. The human genome DNA encodes all life activities of human, so theoretically, the risk of a detected object suffering from a certain disease can be predicted at an early stage by detecting the genome DNA, and even the detected object can be predicted at birth.
The development of tumors is a very complex process. The molecular genetics basis of tumor research is expected to provide a simple and feasible method for common detection, clinical diagnosis, personalized treatment, disease tracking after recovery and the like. However, the individual differences of patients, the intercrossing of related biomolecular events at different stages of development, etc. all bring great difficulties to the work. It is clearly impossible and not scientific to use single molecular genetic changes to diagnose tumors. The inventor applies modern molecular biology technology to carry out combined analysis on a plurality of STRs of genomic DNA of a detected object, and combines statistical analysis methods such as discriminant analysis and the like, thereby inventing a kit for early warning of female breast malignant tumor susceptibility.
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Fig. 1 is a schematic diagram of modules included in a data processing and determining device in the system for predicting susceptibility to female breast malignant tumor according to the present invention.
Detailed Description
The invention will be better understood from the following description of specific embodiments thereof, taken in conjunction with the accompanying drawings and examples. The experimental procedures used in the following examples are all conventional procedures unless otherwise specified. Reference will now be made in detail to embodiments of the present invention, examples of which are illustrated in the accompanying drawings, wherein like or similar reference numerals refer to the same or similar elements or elements having the same or similar function throughout. The embodiments described below with reference to the accompanying drawings are illustrative only for the purpose of explaining the present invention, and are not to be construed as limiting the present invention.
The PCR amplification apparatus in the examples was a Mastercycler nexus amplification apparatus (purchased from eppendorf, USA);
the STR locus fragment analyzer in the examples was a 3730XL sequencing analyzer (purchased from ABI, usa);
the DNA extraction kit in the examples was a blood DNAout kit (purchased from engze, beijing);
the rotational speed of all the centrifuges in the examples was 3000 g/min.
Example 1A system for predicting susceptibility to female breast cancer
A system for predicting susceptibility to a female breast malignancy, comprising:
a device for obtaining the repeat times of the STR locus short tandem sequence of the sample DNA;
data processing and decision device, comprising the following modules (fig. 1):
the data input module is used for inputting the age, the sex and the STR locus short tandem sequence repetition times of the detected object;
the database management module is used for the operation management of data storage, modification, deletion, inquiry and printing;
the data calculation module is used for calculating a discrimination function result according to the repeat times of the STR locus short serial sequence in the data input module;
and the analysis, discrimination and result output module is used for comparing the discrimination function results so as to predict the susceptibility of the female breast malignant tumor and output the result.
Wherein the content of the first and second substances,
the number of times of the STR locus short tandem sequence repetition is 6 pairs of the number of times of the STR locus short tandem sequence repetition:
the discriminant function includes:
first discriminant function FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160
Second discrimination function FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413
In the case of the discriminant function,
X1the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-1;
X2is the larger fragment of the two alleles of STR-1The number of repetitions of the short tandem sequence of (a);
X3the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-2;
X4the number of repeats of the short tandem sequence for the larger of the two alleles of STR-2;
X5the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-3;
X6the number of repeats of the short tandem sequence for the larger of the two alleles of STR-3;
X7the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-4;
X8the number of repeats of the short tandem sequence for the larger of the two alleles of STR-4;
X9the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-5;
X10the number of repeats of the short tandem sequence for the larger of the two alleles of STR-5;
X11the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-6;
X12the number of repeats of the short tandem sequence for the larger fragment of both alleles of STR-6.
Wherein, X1 -X12Calculated from the fragment length and the following formula, where round stands for rounded integer:
X1＝round[(L1-191)/3]；X2＝round[(L2-191)/3]；
X3＝round(L3-379)；X4＝round(L4-379)；
X5＝round[(L5-202)/2]；X6＝round[(L6-202)/2]；
X7＝round[(L7-359)/2]；X8＝round[(L8-359)/2]；
X9＝round[(L9-278)/2]；X10＝round[(L10-278)/2]；
X11＝round[(L11-200)/4]；X12＝round[(L12-200)/4]；
X1 -X12in, L1Is the smaller segment length value, L, of the two alleles of STR-12Is the larger fragment length value of the two alleles of STR-1;
L3is the smaller segment length value, L, of the two alleles of STR-24Is the larger fragment length value of the two alleles of STR-2;
L5is the smaller segment length value, L, of the two alleles of STR-36Is the larger fragment length value of the two alleles of STR-3;
L7is the smaller segment length value, L, of the two alleles of STR-48Is the larger fragment length value of the two alleles of STR-4;
L9is the smaller segment length value, L, of the two alleles of STR-510Is the larger fragment length value of the two alleles of STR-5;
L11is the smaller segment length value, L, of the two STR-6 alleles12Is the larger fragment length value of the two alleles of STR-6;
the analysis discrimination and result output module outputs a first discrimination function FBCAnd a second discrimination function FBNIf F is the result of the calculation ofBC>FBNOutputting a prediction result that the probability of the female breast malignant tumor of the detected object is more than or equal to 90.0%; if FBC≤FBNAnd outputting a prediction result that the probability that the detected object does not suffer from the female breast malignant tumor is more than or equal to 77.8 percent.
Example 2A kit for predicting susceptibility to female breast cancer
A kit for predicting susceptibility to female breast malignant tumor comprises the following components: STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer, STR-6 primer, PCR amplification reaction liquid, LIZ-500 molecular weight internal standard, deionized formamide and an instruction book.
The concentrations of the STR-1 primer, the STR-2 primer, the STR-3 primer, the STR-4 primer, the STR-5 primer and the STR-6 primer are all 10 mu M, and the primer sequences are shown in the following table:
the PCR amplification reaction solution is a mixed solution of the following reagents: TaqDNA polymerase (5U/. mu.L), Tris-HCl (100mM, pH 8.8 at 25 ℃), KCl (500mM), ethylphenylpolyethyleneglycol (0.8% (v/v)), MgCl2(25mM), dNTP (10mM), deionized water.
Storing the PCR amplification reaction solution at-20 ℃; LIZ-500 molecular weight internal standard is preserved at-20 ℃; storing deionized formamide at 2-8 deg.C.
The kit further comprises instructions for use.
Example 3 Using the System of example 1 and the kit of example 2 to predict the risk of developing a female breast malignancy in the subject
The detected object is: women, age 52, visit the mammary surgery department of Jilin university's second hospital, with full informed examination purpose and use, under his voluntary premise, signed an informed consent, and collected 1mL of anticoagulated blood via the peripheral vein.
The following procedure was carried out using the kit of example 2 according to the method described in the kit instructions:
(1) extracting sample DNA: extracting blood genome DNA by using a DNA extraction kit;
(2) PCR reaction
(2-1) taking out the STR-1 primer, the STR-2 primer, the STR-3 primer, the STR-4 primer, the STR-5 primer, the STR-6 primer and the PCR amplification reaction solution from a refrigerator, balancing to room temperature, fully dissolving each component, and respectively and rapidly centrifuging for 10 seconds;
(2-2) adding 100ng of sample DNA into 60 mu L of PCR amplification reaction solution, adding deionized water to supplement to 115.2 mu L, fully and uniformly mixing, quickly centrifuging for 10 seconds, and subpackaging the mixed solution into 6 PCR reaction tubes according to 19.2 mu L/hole;
(2-3) respectively adding an STR-1 primer, an STR-2 primer, an STR-3 primer, an STR-4 primer, an STR-5 primer and an STR-6 primer into the 6 PCR reaction tubes in the step (2-2) according to 0.8 mu L/hole; covering a PCR reaction tube cover, recording the sample adding condition, quickly centrifuging for 10 seconds, then transferring the PCR reaction tube to a corresponding position of a sample groove of a PCR amplification instrument, recording the placing sequence, and starting the PCR amplification reaction; the amplification reaction conditions are as follows: 3 minutes at 95 ℃; 30 seconds at 95 ℃, 30 seconds at 60 ℃ and 30 seconds at 72 ℃ for 10 cycles; 30 seconds at 95 ℃, 30 seconds at 55 ℃, 30 seconds at 72 ℃ and 20 cycles; 6 groups of PCR amplification products are obtained at 72 ℃ for 6 minutes;
(3) STR fragment analysis
(3-1) adding 990 mu L of deionized formamide into 10 mu L of LIZ-500 molecular weight internal standard, fully and uniformly mixing, quickly centrifuging for 10 seconds, respectively adding into a sequencing reaction tube according to 10 mu L/hole, and quickly centrifuging for 10 seconds;
(3-2) adding the 6 groups of PCR amplification products into 6 sequencing reaction tubes according to 1 mu L/hole respectively, and quickly centrifuging for 10 seconds; then transferring the sequencing reaction tube to a corresponding position of a sample tank of a PCR (polymerase chain reaction) amplification instrument, heating at 98 ℃ for 5 minutes, immediately placing the sequencing reaction tube on an ice-water mixture after the program is finished, rapidly cooling to 0 ℃, and rapidly centrifuging for 10 seconds; then transferring the sequencing reaction tube to a corresponding position of a sample groove of an STR locus fragment analyzer, recording the placement sequence, and performing fragment analysis detection;
(4) analysis and determination of results
(4-1) respectively recording the fragment lengths of two alleles at each site of STR-1, STR-2, STR-3, STR-4, STR-5 and STR-6 according to the fragment analysis result: the length of the smaller of the two STR-1 alleles is recorded as L1And the length of the larger fragment of the two STR-1 alleles is designated as L2(ii) a The length of the smaller fragment of the two STR-2 alleles is recorded as L3The larger of the two alleles of STR-2Segment length value is noted as L4(ii) a The length of the smaller fragment of the two STR-3 alleles is recorded as L5And the length of the larger fragment of the two STR-3 alleles is marked as L6(ii) a The length of the smaller fragment of the two STR-4 alleles is recorded as L7And the length of the larger fragment of the two STR-4 alleles is marked as L8(ii) a The length of the smaller of the two STR-5 alleles is recorded as L9And the length of the larger fragment of the two STR-5 alleles is designated as L10(ii) a The length of the smaller fragment of the two STR-6 alleles was designated L11And the length of the larger fragment of the two STR-6 alleles is marked as L12(ii) a The results show that: l is1＝273.59，L2＝276.27，L3＝403.24，L4＝403.24，L5＝229.33，L6＝246.78，L7＝384.99，L8＝396.6，L9＝312.13，L10＝320.45，L11＝252.54，L12＝264.19。
(4-2) the length of the fragment is calculated from the following formula, and is denoted as X1-X12Where round stands for rounded integer:
X1＝round[(L1-191)/3]＝28；X2＝round[(L2-191)/3]＝28；
X3＝round(L3-379)＝24；X4＝round(L4-379)＝24；
X5＝round[(L5-202)/2]＝14；X6＝round[(L6-202)/2]＝22；
X7＝round[(L7-359)/2]＝13；X8＝round[(L8-359)/2]＝19；
X9＝round[(L9-278)/2]＝17；X10＝round[(L10-278)/2]＝21；
X11＝round[(L11-200)/4]＝13；X12＝round[(L12-200)/4]＝16。
(4-3) predicting susceptibility of the subject to the female breast malignancy using a computer running the system for predicting susceptibility to female breast malignancy described in example 1:
inputting the age, the sex and the STR locus short tandem sequence repetition times of the detected object into a system through a data input module, and calculating the result of a discriminant function through a data calculation module:
first discriminant function FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160＝1778.095
Second discrimination function FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413＝1773.798
Analyzed, determined and result output module compared FBCValue sum FBNValue, FBC>FBNAnd outputting a prediction result that the probability of the female breast malignant tumor of the detected object is more than or equal to 90.0 percent.
The examinee is subjected to breast cancer improvement radical operation after seeing a doctor, the pathological examination confirms that the examinee is the breast invasive ductal carcinoma, and the clinical diagnosis result of the examinee is consistent with the prediction result of the kit.
Example 4 Using the System of example 1 and the kit of example 2 to predict the risk of developing a female breast malignancy in the subject
The detected object is: the female, 60 years old, in the second hospital of Jilin university, the surgery of mammary gland, conduct mammary duct endoscopy, in the full notice of the purpose and use of the examination, under the premise of its voluntary, sign the informed consent, and collect 1mL of anticoagulation through the peripheral vein.
The same treatments and tests were carried out on blood samples, with reference to the prediction method of example 3, and the results show that: l is1＝265.48，L2＝270.86，L3＝404.17，L4＝404.17，L5＝228.62，L6＝228.62，L7＝398.52，L8＝404.64，L9＝324.6，L10＝330.99，L11＝255.66，L12＝263.75。
Calculated according to the fragment length and the following formula, denoted X1-X12Where round stands for rounded integer:
X1＝round[(L1-191)/3]＝25；X2＝round[(L2-191)/3]＝27；
X3＝round(L3-379)＝25；X4＝round(L4-379)＝25；
X5＝round[(L5-202)/2]＝13；X6＝round[(L6-202)/2]＝13；
X7＝round[(L7-359)/2]＝20；X8＝round[(L8-359)/2]＝23；
X9＝round[(L9-278)/2]＝23；X10＝round[(L10-278)/2]＝26；
X11＝round[(L11-200)/4]＝14；X12＝round[(L12-200)/4]＝16。
using a computer running the system for predicting susceptibility to female breast malignancies described in example 1, a susceptibility prediction of subjects to female breast malignancies is made:
inputting the age, the sex and the STR locus short tandem sequence repetition times of the detected object into a system through a data input module, and calculating the result of a discriminant function through a data calculation module:
first discriminant function FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160＝1900.929
Second discrimination function FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413＝1903.959
Analyzed, determined and result output module compared FBCValue sum FBNValue, FBC≤FBNAnd outputting a prediction result that the probability that the detected object does not suffer from the female breast malignant tumor is more than or equal to 77.8 percent.
The detected object is diagnosed as the papillary tumor in the mammary duct after the diagnosis, which belongs to benign tumor, and the clinical diagnosis result of the detected object is consistent with the prediction result of the kit.
Example 5 Using the System of example 1 and the kit of example 2 to predict the risk of developing a female breast malignancy in the subject
The detected object is: the female, 59 years old, in the second hospital of Jilin university, of mammary surgery, underwent a radical improvement surgery for breast cancer, signed an informed consent on the premise of fully informing the purpose and use of examination, and collected 1mL of anticoagulation blood via the peripheral vein.
The same treatments and tests were carried out on blood samples, with reference to the prediction method of example 3, and the results show that: l is1＝268.46，L2＝287.38，L3＝403.19，L4＝403.19，L5＝229.08，L6＝229.08，L7＝383.05，L8＝388.80，L9＝312.05，L10＝318.12，L11＝253.32，L12＝260.50。
Calculated according to the fragment length and the following formula, denoted X1-X12Where round stands for rounded integer:
X1＝round[(L1-191)/3]＝23；X2＝round[(L2-191)/3]＝28；
X3＝round(L3-379)＝24；X4＝round(L4-379)＝24；
X5＝round[(L5-202)/2]＝21；X6＝round[(L6-202)/2]＝22；
X7＝round[(L7-359)/2]＝12；X8＝round[(L8-359)/2]＝20；
X9＝round[(L9-278)/2]＝16；X10＝round[(L10-278)/2]＝17；
X11＝round[(L11-200)/4]＝13；X12＝round[(L12-200)/4]＝13；
using a computer running the system for predicting susceptibility to female breast malignancies described in example 1, a susceptibility prediction of subjects to female breast malignancies is made:
inputting the age, the sex and the STR locus short tandem sequence repetition times of the detected object into a system through a data input module, and calculating the result of a discriminant function through a data calculation module:
first discriminant function FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160＝1745.99
Second discrimination function FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413＝1742.763
Analyzed, determined and result output module compared FBCValue sum FBNValue, FBC>FBNAnd outputting a prediction result that the probability of the female breast malignant tumor of the detected object is more than or equal to 90.0 percent.
The examined person is diagnosed with the breast invasive ductal carcinoma, and the clinical diagnosis result of the examined person is consistent with the prediction result of the kit.
Example 6 prediction of the risk of acquiring a female breast malignancy in a subject using the system of example 1 and the kit of example 2
The detected object is: women, age 33, visiting the department of mammary surgery at the second hospital, Jilin university, underwent mastectomy, with full informed examination and use, signed informed consent and collected anticoagulated 1mL via the peripheral vein on the premise of their own accord.
The same treatments and tests were carried out on blood samples, with reference to the prediction method of example 3, and the results show that: l is1＝265.37，L2＝276.26，L3＝404.10，L4＝404.10，L5＝228.72，L6＝230.87，L7＝388.82，L8＝388.82，L9＝324.57，L10＝324.57，L11＝262.90，L12＝267.41。
Calculated according to the fragment length and the following formula, denoted X1-X12Where round stands for rounded integer:
X1＝round[(L1-191)/3]＝25；X2＝round[(L2-191)/3]＝28；
X3＝round(L3-379)＝25；X4＝round(L4-379)＝25；
X5＝round[(L5-202)/2]＝13；X6＝round[(L6-202)/2]＝14；
X7＝round[(L7-359)/2]＝15；X8＝round[(L8-359)/2]＝15；
X9＝round[(L9-278)/2]＝23；X10＝round[(L10-278)/2]＝23；
X11＝round[(L11-200)/4]＝16；X12＝round[(L12-200)/4]＝17。
using a computer running the system for predicting susceptibility to female breast malignancies described in example 1, a susceptibility prediction of subjects to female breast malignancies is made:
inputting the age, the sex and the STR locus short tandem sequence repetition times of the detected object into a system through a data input module, and calculating the result of a discriminant function through a data calculation module:
first discriminant function FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160＝1901.877
Second discrimination function FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413＝1905.21
Analyzed, determined and result output module compared FBCValue sum FBNValue, FBC≤FBNAnd outputting a prediction result that the probability that the detected object does not suffer from the female breast malignant tumor is more than or equal to 77.8 percent.
The subject is diagnosed as the fibroadenoma of breast after the visit, belongs to benign tumor, and the clinical diagnosis result of the subject is consistent with the prediction result of the kit.
The foregoing is a preferred embodiment of the present invention and is not intended to limit the present invention, and it should be understood that any changes, modifications, substitutions and alterations (e.g., addition, subtraction, change of STR sites, use of cells or tissues from other sources, use of other statistical methods, etc.) made without departing from the principles and spirit of the present invention are intended to be included within the scope of the present invention.
SEQUENCE LISTING
<110> Jilin university
<120> female breast malignant tumor susceptibility prediction kit and system
<130> DI18-8180-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)
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tgccccatag gttttgaact 20
Claims (7)
1. A kit for predicting susceptibility to female breast malignant tumor comprises the following components: STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer and STR-6 primer, wherein the STR-1 primer, STR-2 primer, STR-3 primer, STR-4 primer, STR-5 primer and STR-6 primer are respectively used for amplifying target fragments containing the following short tandem sequences so as to determine the repetition times of the following short tandem sequences:
The sequences of the STR-1 primer, the STR-2 primer, the STR-3 primer, the STR-4 primer, the STR-5 primer and the STR-6 primer are as follows:
wherein, the kit further comprises: PCR amplification reaction liquid, LIZ-500 molecular weight internal standard, deionized formamide and an instruction book,
the application instruction records a use method of the female breast malignant tumor susceptibility prediction kit, which comprises the following steps:
(1) extracting sample DNA;
(2) PCR reaction
(2-1) taking out the STR-1 primer, the STR-2 primer, the STR-3 primer, the STR-4 primer, the STR-5 primer, the STR-6 primer and the PCR amplification reaction solution from a refrigerator, balancing to room temperature, fully dissolving each component, and respectively and rapidly centrifuging for 10 seconds;
(2-2) adding 30-300ng of sample DNA into 60 mu L of PCR amplification reaction solution, adding deionized water to supplement to 115.2 mu L, fully and uniformly mixing, quickly centrifuging for 10 seconds, and subpackaging the mixed solution into 6 PCR reaction tubes according to 19.2 mu L/hole;
(2-3) respectively adding an STR-1 primer, an STR-2 primer, an STR-3 primer, an STR-4 primer, an STR-5 primer and an STR-6 primer into the 6 PCR reaction tubes in the step (2-2) according to 0.8 mu L/hole; covering a PCR reaction tube cover, recording the sample adding condition, quickly centrifuging for 10 seconds, then transferring the PCR reaction tube to a corresponding position of a sample groove of a PCR amplification instrument, recording the placing sequence, and starting the PCR amplification reaction; the amplification reaction conditions are as follows: 3 minutes at 95 ℃; 30 seconds at 95 ℃, 30 seconds at 60 ℃ and 30 seconds at 72 ℃ for 10 cycles; 30 seconds at 95 ℃, 30 seconds at 55 ℃, 30 seconds at 72 ℃ and 20 cycles; 6 groups of PCR amplification products are obtained at 72 ℃ for 6 minutes;
(3) STR fragment analysis
(3-1) adding 990 mu L of deionized formamide into 10 mu L of LIZ-500 molecular weight internal standard, fully and uniformly mixing, quickly centrifuging for 10 seconds, respectively adding into a sequencing reaction tube according to 10 mu L/hole, and quickly centrifuging for 10 seconds;
(3-2) adding the 6 groups of PCR amplification products into 6 sequencing reaction tubes according to 1 mu L/hole respectively, and quickly centrifuging for 10 seconds; then transferring the sequencing reaction tube to a corresponding position of a sample tank of a PCR (polymerase chain reaction) amplification instrument, heating at 98 ℃ for 5 minutes, immediately placing the sequencing reaction tube on an ice-water mixture after the program is finished, rapidly cooling to 0 ℃, and rapidly centrifuging for 10 seconds; then transferring the sequencing reaction tube to a corresponding position of a sample groove of an STR locus fragment analyzer, recording the placement sequence, and performing fragment analysis detection;
(4) analysis and determination of results
(4-1) respectively recording the fragment lengths of two alleles at each site of STR-1, STR-2, STR-3, STR-4, STR-5 and STR-6 according to the fragment analysis result:
the length of the smaller of the two STR-1 alleles is recorded as L1And the length of the larger fragment of the two STR-1 alleles is designated as L2；
The length of the smaller fragment of the two STR-2 alleles is recorded as L3And the length of the larger fragment of the two STR-2 alleles is designated as L4；
The length of the smaller fragment of the two STR-3 alleles is recorded as L5And the length of the larger fragment of the two STR-3 alleles is marked as L6；
The length of the smaller fragment of the two STR-4 alleles is recorded as L7And the length of the larger fragment of the two STR-4 alleles is marked as L8；
The length of the smaller of the two STR-5 alleles is recorded as L9And the length of the larger fragment of the two STR-5 alleles is designated as L10；
The length of the smaller fragment of the two STR-6 alleles was designated L11And the length of the larger fragment of the two STR-6 alleles is marked as L12；
(4-2) the number of repetitions of the short tandem sequence is calculated from the fragment length and the following formula, and is denoted as X1-X12Where round stands for rounded integer:
X1＝round[(L1-191)/3]；X2＝round[(L2-191)/3]；
X3＝round(L3-379)；X4＝round(L4-379)；
X5＝round[(L5-202)/2]；X6＝round[(L6-202)/2]；
X7＝round[(L7-359)/2]；X8＝round[(L8-359)/2]；
X9＝round[(L9-278)/2]；X10＝round[(L10-278)/2]；
X11＝round[(L11-200)/4]；X12＝round[(L12-200)/4]；
(4-3) substituting the number of the short tandem sequence repetitions into a preset discriminant function:
FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160
FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413；
(4-4) prediction of susceptibility to female breast malignancy:
comparison FBCValue sum FBNValue if FBC>FBNThen the probability of the female breast malignant tumor of the detected object is predicted to be more than or equal to 90.0 percent; if FBC≤FBNAnd predicting that the probability that the detected object does not suffer from the female breast malignant tumor is more than or equal to 77.8 percent.
2. The kit for predicting susceptibility to breast malignancy in women according to claim 1, wherein: the using concentration of the STR-1 primer, the STR-2 primer, the STR-3 primer, the STR-4 primer, the STR-5 primer and the STR-6 primer is 10 mu M.
3. The kit for predicting susceptibility to breast malignancy in women according to claim 1, wherein: the PCR amplification reaction solution is a mixed solution of the following reagents: TaqDNA polymerase 5U/. mu. L, Tris-HCl 100mM, KCl 500mM, ethylphenylpolyethylene glycol 0.8 vol%, MgCl225mM, dNTP 10mM and deionized water; wherein Tris-HCl has a pH of 8.8 at 25 ℃.
4. The kit for predicting susceptibility to breast malignancy in women according to claim 1, wherein: the sample is whole blood of a subject.
5. Use of the kit for predicting susceptibility to female breast cancer according to any one of claims 1 to 4 in the preparation of a diagnostic product for female breast cancer.
6. A system for predicting susceptibility to a female breast malignancy, comprising:
means for obtaining the number of repetitions of the following short tandem STR loci of the sample DNA:
；
Data processing and decision device, comprising the following modules:
the data input module is used for inputting the age, the sex and the STR locus short tandem sequence repetition times of the detected object;
the database management module is used for the operation management of data storage, modification, deletion, inquiry and printing;
the data calculation module is used for calculating a discrimination function result according to the repeat times of the STR locus short serial sequence in the data input module;
an analysis, discrimination and result output module for comparing the discrimination function results to make the female breast malignant tumor susceptibility prediction and outputting the result,
wherein the number of times X of repetition of STR site short tandem sequences of sample DNA obtained using the kit of any one of claims 1 to 51-X12(ii) a And
wherein:
the discriminant function includes:
first discriminant function FBC＝12.108X1-0.933X2+15.490X3+112.846X4+2.606X5+0.178X6+6.318X7-1.147X8+2.405X9-4.504X10+2.549X11+5.507X12-1783.160
Second discrimination function FBN＝12.504X1-1.212X2+15.869X3+115.560X4+2.561X5-0.026X6+6.887X7-1.615X8+2.619X9-4.518X10+2.486X11+5.478X12-1860.413
In the case of the discriminant function,
X1the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-1;
X2the number of repeats of the short tandem sequence for the larger of the two alleles of STR-1;
X3the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-2;
X4the number of repeats of the short tandem sequence for the larger of the two alleles of STR-2;
X5the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-3;
X6the number of repeats of the short tandem sequence for the larger of the two alleles of STR-3;
X7the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-4;
X8the number of repeats of the short tandem sequence for the larger of the two alleles of STR-4;
X9the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-5;
X10the number of repeats of the short tandem sequence for the larger of the two alleles of STR-5;
X11the number of repeats of the short tandem sequence for the smaller of the two alleles of STR-6;
X12the number of repeats of the short tandem sequence for the larger of the two alleles of STR-6;
wherein, X1 -X12Calculated from the fragment length and the following, where round stands for rounded integer:
X1＝round[(L1-191)/3]；X2＝round[(L2-191)/3]；
X3＝round(L3-379)；X4＝round(L4-379)；
X5＝round[(L5-202)/2]；X6＝round[(L6-202)/2]；
X7＝round[(L7-359)/2]；X8＝round[(L8-359)/2]；
X9＝round[(L9-278)/2]；X10＝round[(L10-278)/2]；
X11＝round[(L11-200)/4]；X12＝round[(L12-200)/4]；
X1 -X12in, L1Is the smaller segment length value, L, of the two alleles of STR-12Is the larger fragment length value of the two alleles of STR-1;
L3is the smaller segment length value, L, of the two alleles of STR-24Is the larger fragment length value of the two alleles of STR-2;
L5is the smaller segment length value, L, of the two alleles of STR-36Is the larger fragment length value of the two alleles of STR-3;
L7is the smaller segment length value, L, of the two alleles of STR-48Is the larger fragment length value of the two alleles of STR-4;
L9is the smaller segment length value, L, of the two alleles of STR-510Is the larger fragment length value of the two alleles of STR-5;
L11is the smaller segment length value, L, of the two STR-6 alleles12Is the larger fragment length value of the two alleles of STR-6;
the analysis discrimination and result output module outputs a first discrimination function FBCAnd a second discrimination function FBNIf F is the result of the calculation ofBC>FBNThen, a prediction node that the probability of the female breast malignant tumor of the detected object is more than or equal to 90.0 percent is outputFruit; if FBC≤FBNAnd outputting a prediction result that the probability that the detected object does not suffer from the female breast malignant tumor is more than or equal to 77.8 percent.
7. The use of the system of claim 6 in the preparation of a product for predicting a malignant tumor of a female breast or a product for diagnosing a malignant tumor of a female breast.
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