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Kit and method for polymorphism detection of metabolism ability genes MTHFR and MTRR of folic acid

A technology of gene polymorphism and detection kits, which is applied in biochemical equipment and methods, microbe measurement/inspection, DNA/RNA fragments, etc. It can solve the problems of high template quality requirements and insufficient timeliness, and achieve system stability , good sensitivity and specificity

Inactive Publication Date: 2020-05-08
重庆浦洛通基因医学研究院有限公司
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  • Claims
  • Application Information

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Problems solved by technology

[0005] The purpose of the present invention is to solve the problem of high template quality requirements and insufficient timeliness in the polymorphism detection of folic acid metabolism ability genes MTHFR and MTRR, and provides a genotyping detection of folate metabolism ability gene polymorphism sites Kits and methods

Method used

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  • Kit and method for polymorphism detection of metabolism ability genes MTHFR and MTRR of folic acid
  • Kit and method for polymorphism detection of metabolism ability genes MTHFR and MTRR of folic acid
  • Kit and method for polymorphism detection of metabolism ability genes MTHFR and MTRR of folic acid

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Embodiment 1

[0070] 1. Primers, probes and inhibitors for detecting polymorphisms of MTHFR gene 677C>T, MTHFR gene 1298A>C and MTRR gene 66A>G

[0071] 1) Design specific arms-PCR primers and probes according to the MTHFR gene sequence (Gene ID: 4524) published in the NCBI database, taking the 677C>T site (rs1801133) of the MTHFR gene as a reference, and referring to Premier's primer primer 5.0. Using the mutant plasmid and wild-type plasmid constructed by genetic engineering as templates, a real-time fluorescent PCR detection system was established to achieve high sensitivity and high specificity detection of rs1801133.

[0072] The primers and probes for detecting the 677C>T mutation of the MTHFR gene are characterized in that the detection primers and probes are designed according to the rs1801133 site of the MTHFR gene, and the sequences are as follows:

[0073] Wild-type ARMS upstream primer, 677-WF: AAGCTGCGTGATGATGACATAGG

[0074] Mutant ARMS upstream primer, 677-MF: AAGCTGCGTGATGA...

Embodiment 2

[0148] The usage of the kit of the present invention will be described in detail below in conjunction with specific example 2. Example 2 is implemented under the technical premise of the present invention, and provides detailed implementation methods and specific operating procedures. In Example 2, 19 blood samples were collected and named as samples No. 1-19 respectively. The results detected by the method of the present invention were compared with the results of first-generation sequencing to confirm the accuracy of the present invention.

[0149] 1. Extraction of Genomic DNA from 19 Blood Samples

[0150] Sample DNA was extracted using a commercially mature small amount of nucleic acid extraction kit. In the present invention, HiPure Blood DNA Midi Kit II (D3113-03, Magen), a nucleic acid extraction kit from Meiji Biology, is used, and the specific usage method refers to the kit instruction manual. The samples to be tested were extracted with commercial kits and stored fo...

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Abstract

The invention provides a kit and method for genotyping detection of polymorphism sites of metabolism ability genes of folic acid. The kit includes a mutation-type reaction system and a wild-type reaction system, wherein the reaction systems include rs1801133 groups, rs1801131 groups and rs1801394 groups, and the structures of primers, probes and inhibitors in each group are disclosed. The problemsof high requirements for template quality and insufficient timeliness can be solved during detection of polymorphism typing of the metabolism genes MTHFR and MTRR of folic acid, only the two tubes ofreaction systems are needed, one-time sampling and one-time closed detection can be realized, and polymorphism of three sites of 677C>T, 1298A>C and 66A>G of the gene MTHFR are detected in real time;and the kit and method have stable systems and good sensitivity and specificity.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a kit and a use method for detecting the polymorphisms of folate metabolism ability genes MTHFR and MTRR. Background technique [0002] Folic acid is an essential substance for cell growth and tissue repair, and is especially important for pregnant women during embryonic development. According to the serum sample survey of women of childbearing age before marriage in my country, 21% and 38% of women in urban and rural areas in my country lack folic acid respectively. According to relevant reports from the Ministry of Health and the Center for Disease Control and Prevention, 900,000 newborns are born with defects and diseases every year in my country. The top five birth defects include: congenital heart disease, neural tube defects, cleft lip and palate, Down syndrome, Multiple fingers (toes), these are related to the lack of folic acid in pregnant women. [0003] Recent ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12N15/11
CPCC12Q1/6883C12Q2600/106C12Q2600/156
Inventor 韩勋领孙松松罗锋
Owner 重庆浦洛通基因医学研究院有限公司
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