Fluorescence quantitative polymerase chain reaction (PCR) detection system for osteosarcoma gene screening, and application of fluorescence quantitative PCR detection system

A fluorescence quantification and detection system technology, applied in the direction of microbial determination/inspection, biochemical equipment and methods, etc., can solve the problems of tumor suppressor gene inactivation, weak site correlation, imbalance of major signaling pathways, etc., to increase the number of physical examinations. Frequency, specificity, and low-cost effects

Inactive Publication Date: 2018-03-02
SHENZHEN MEIGENE MEDICAL LAB
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Problems solved by technology

Moreover, existing data also show that there are a large number of genetic and molecular changes in osteosarcoma, including the inactivation of tumor suppressor genes caused by chromosome gain, loss or rearrangement of chromosome regions, and the imbalance of major signaling pathways, etc.
[0010] The existing genetic loci related to osteosarcoma are strong association loci based on the large-sample GWAS study of European populations. When applied to Asian populations, especially Chinese populations, the loci are weakly correlated and difficult to obtain through detection. More accurate results, so it is necessary to establish an evaluation system for the Chinese population

Method used

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  • Fluorescence quantitative polymerase chain reaction (PCR) detection system for osteosarcoma gene screening, and application of fluorescence quantitative PCR detection system
  • Fluorescence quantitative polymerase chain reaction (PCR) detection system for osteosarcoma gene screening, and application of fluorescence quantitative PCR detection system
  • Fluorescence quantitative polymerase chain reaction (PCR) detection system for osteosarcoma gene screening, and application of fluorescence quantitative PCR detection system

Examples

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Effect test

Embodiment 1

[0053] In this example, an osteosarcoma fluorescent quantitative PCR detection kit was used to detect osteosarcoma-related genes, in which five polymorphic sites on five genes related to osteosarcoma were detected, including rs231775, rs1129055, rs7034162, and rs1906953 , rs7591996 loci, a total of 5 loci. Using specifically designed primers and probes, the amplification system and conditions are optimized, and the detection of two polymorphic sites on two genes is completed simultaneously in the same system.

[0054] In this example, a single fluorescently labeled probe, Roche 480 platform, and finally detected rs231775, rs1129055, rs7034162, rs1906953, rs7591996 sites by melting curve peak pattern.

[0055] Specifically, in the dual fluorescence quantitative PCR detection method for rs231775, rs1129055, rs7034162, rs1906953, and rs7591996 sites, the primers and probe designs for rs231775, rs1129055, rs7034162, rs1906953, and rs7591996 sites are shown in Table 1 below.

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Embodiment 2

[0080] The five loci in Example 1 are from the articles published by Bezzina et al. and Dr. Ar-king respectively. The article obtained two susceptibility loci directly related to osteosarcoma through GWAS research, and calculated the two loci The risk value of the risk allele type, that is, the OR value.

[0081] GWAS studies have found that common genetic variations in NFIB, GRM4 and 2p25.2 may be associated with osteosarcoma. Mirabello L et al found that the SNP site (rs7034162) on the NFIB gene is a risk factor for osteosarcoma cell migration, proliferation and colony formation. The rs7034162 mutation (genetic variation in the germline) is a susceptibility gene for tumor cell metastasis.

[0082] Osteosarcoma is the most common primary bone malignancy in adolescents and young adults. To better understand the genetic etiology of osteosarcoma, Savage S A et al conducted a multistage genome-wide association study including 941 osteosarcoma patients and 3291 European adult co...

Embodiment 3

[0104] The method of the present invention extracts DNA by collecting the genetic sample of subject then, detects with Roche LightCycler480 real-time quantitative instrument, and the result obtained is as follows: figure 1 , 3, 5, 7, 9.

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Abstract

The invention relates to the technical field of biological detection, in particular to a fluorescence quantitative polymerase chain reaction (PCR) detection kit, and further discloses application of the fluorescence quantitative PCR detection kit in detection of osteosarcoma. The fluorescence quantitative PCR detection system comprises a detection kit for genes at specific loci, calculates the score of osteosarcoma risk according to risk values of the osteosarcoma risk at all the loci, and performs crowd classification according to the existing database; the osteosarcoma risk level of an individual is obtained according to the individual score, so that the osteosarcoma risk of a person subjected to detection can be evaluated, the people with high risk is guided to improve the lifestyle, inducing factors are avoided, and the frequency of physical examination is increased. Furthermore, the fluorescence quantitative PCR detection system lays a foundation for the evaluation of the specificosteosarcoma of Chinese population by establishing a Chinese population database for the five selected gene polymorphic loci.

Description

technical field [0001] The invention relates to the technical field of biological detection, in particular to a fluorescent quantitative PCR detection kit, and further discloses its application for detecting osteosarcoma. Background technique [0002] With the continuous development of scientific research, especially after the completion of the Human Genome Project and the full development of the post-genome project, the application of the latest technology dominated by genetic testing in the field of disease prevention and control has attracted widespread attention. At present, preliminary progress has been made in the prediction of disease risk using genetic theories and methods. Commonly used genetic testing methods include: direct sequencing (DS), ligase detection reaction (LDR), restriction Fragment length polymorphism analysis (restriction fragment length polymorphism, RFLP), denaturing high performance liquid chromatography (denaturing high performance liquid chromato...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6851
CPCC12Q1/6851C12Q2563/107C12Q2531/113C12Q2537/165
Inventor 吴少鸿周文根李欣姜萍萍黄君任飞詹延延赵世林李政
Owner SHENZHEN MEIGENE MEDICAL LAB
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