Human MTHFR and MTRR gene polymorphism detection primer, probe, test kit and method

A gene polymorphism, detection kit technology, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc. and other problems, to achieve the effect of rapid detection, avoid cross-contamination of samples, and reliable results

Active Publication Date: 2017-04-26
广州达晖生物技术股份有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the sequencing method is the gold standard for the detection of genetic polymorphisms, the operation is cumbersome, the workload is heavy, and the instrumen

Method used

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  • Human MTHFR and MTRR gene polymorphism detection primer, probe, test kit and method
  • Human MTHFR and MTRR gene polymorphism detection primer, probe, test kit and method
  • Human MTHFR and MTRR gene polymorphism detection primer, probe, test kit and method

Examples

Experimental program
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Effect test

Embodiment 1

[0047] Example 1 Human MTHFR and MTRR gene polymorphism detection kit

[0048] The test kit described in this embodiment includes the following components:

[0049] Public primers and probes: public primers UP1 (nucleotide sequence shown in SEQ ID NO.1) and UP2 (nucleotide sequence shown in SEQ ID NO.2), beta globin specific upstream primer globinF (SEQ ID NO.3 Nucleotide sequence shown), beta globin specific downstream primer globinR (nucleotide sequence shown in SEQ ID NO.4), beta globin specific probe globin probe (nucleotide sequence shown in SEQ ID NO.5) ;

[0050] Specific Primers and Probes:

[0051] MTHFR677 specific upstream primer MTHFR677F (nucleotide sequence shown in SEQ ID NO.6), MTHFR677 downstream primer MTHFR677R (nucleotide sequence shown in SEQ ID NO.7) for the qualitative detection of MTHFR677C>T gene polymorphism, MTHFR677C specific probe MTHFR677Cprobe (nucleotide sequence shown in SEQ ID NO.8), MTHFR677T specific probe MTHFR677Tprobe (nucleotide seque...

Embodiment 2

[0062] Example 2 Human MTHFR and MTRR gene polymorphism detection method

[0063] Use the test kit of embodiment 1 to detect, comprise the following steps:

[0064] 1. Preparation of DNA

[0065] 30 people were randomly selected to draw peripheral blood and extract DNA according to medical routine with informed consent. The kit of the present invention has no specified requirements on the extraction method of human genomic DNA, and generally, the conventional laboratory method (phenol-chloroform extraction method) or the kit can be used to extract human genomic DNA. Take an appropriate amount of EDTA-K2 or sodium citrate anticoagulated human peripheral whole blood and extract human genomic DNA. The extracted DNA samples must meet the following requirements.

[0066] 1.1 The sample source of the kit of the present invention is human peripheral whole blood genomic DNA, the DNA concentration is 10-50 ng / μL, and the purity A260 / A280 is between 1.8-2.0.

[0067] 1.2 Sample stor...

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Abstract

The invention discloses a human MTHFR and MTRR gene polymorphism detection primer, probe, test kit and method and belongs to the technical field of in-vitro nucleic acid detection. The two most common single nucleotide polymorphisms 677C>T and 1298A>C of the MTHFR and the common mutation site 66A>G of the MTHFR are designed for qualitative detection, and a specific primer containing reference genes beta globin and a probe are designed. The test kit includes a detection primer and probe combination, a reference substance, a PCR reaction liquid and the like. Three independent multiplex PCR reactions are conducted for qualitative detection for the three gene polymorphism sites, namely, MTHFR677C>T, MTHFR1298A>C, and MTHFR 66A>G respectively. The detection primer, the probe, the test kit and the method are strong in specificity, high in sensitivity, simple, rapid and convenient for large-scale application and popularization.

Description

technical field [0001] The invention belongs to the technical field of in vitro nucleic acid detection, and specifically relates to a primer, a probe, a kit and a method for detecting human MTHFR and MTRR gene polymorphisms. Background technique [0002] Folic acid (folic acid) belongs to the B vitamins, is an essential substance for cell growth and reproduction, and participates in many important metabolic reactions of the body. Once folic acid deficiency or folic acid metabolic enzyme deficiency occurs in the body, it will lead to abnormal cell cycle, abnormal DNA and protein methylation reactions, abnormal synthesis of DNA bases and other abnormal biochemical reactions, which will directly or indirectly lead to newborns. Neural tube defects in children or fetuses and cardiovascular disease in adults. At present, it has been confirmed that the genes closely related to folate metabolism include 5,10-methylenetetrahydrofolate reductase (MTHFR) and methionine synthase reduct...

Claims

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Application Information

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IPC IPC(8): C12Q1/68C12N15/11
CPCC12Q1/6858C12Q2537/143C12Q2545/101C12Q2563/107
Inventor 李晓辉徐素茹杨祖婷
Owner 广州达晖生物技术股份有限公司
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