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Detection kit and detection method for direct-expanded type MTHFR and/or MTRR and MTR gene polymorphisms

A gene polymorphism, detection kit technology, applied in biochemical equipment and methods, microbial determination/inspection, DNA/RNA fragments, etc., can solve problems such as unreported detection methods, complex nucleic acid extraction and purification steps, etc. , to prevent foreign imported cross-infection, save experimental consumables and labor costs, and improve experimental efficiency.

Pending Publication Date: 2019-04-12
上海酷乐生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] The above mainstream methods still cannot solve the following two problems: 1) saliva or blood samples require complex nucleic acid extraction and purification steps; 2) multi-point SNP typing detection in one tube
[0009] At present, there is no report on a detection kit and detection method for a one-tube direct expansion type MTHFR and / or MTRR and MTR gene polymorphism

Method used

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  • Detection kit and detection method for direct-expanded type MTHFR and/or MTRR and MTR gene polymorphisms
  • Detection kit and detection method for direct-expanded type MTHFR and/or MTRR and MTR gene polymorphisms
  • Detection kit and detection method for direct-expanded type MTHFR and/or MTRR and MTR gene polymorphisms

Examples

Experimental program
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Effect test

Embodiment 1

[0071] This embodiment is a one-tube direct expansion type MTHFR and / or MTRR and MTR gene polymorphism detection kit, including: primer set and probe set, MTHFR C677T and A1298C joint detection reaction solution, MTRR A66G and MTR A2756G joint detection reaction solution, positive control substance, negative control substance;

[0072] Wherein, the primer set and probe set are: SEQ ID NO.1 to SEQ ID NO.4 for MTHFR C677T site; SEQ ID NO.5 to SEQ ID NO.8 for MTHFR A1298C site; for MTRR A66G SEQ ID NO.9 to SEQ ID NO.12 for the site; SEQ ID NO.13 to SEQ ID NO.16 for the MTR A2756G site, the specific sequences of the above primers and probes are shown in Table 1 above.

[0073] Wherein, the reaction system of the combined detection reaction solution of MTHFR C677T and A1298C includes: improved HotStar Taq enzyme, direct amplification PCR amplification buffer, direct amplification and stabilization reagents, and corresponding primers and probes; the MTRRA66G and MTR A2756G The reac...

Embodiment 2

[0077] This embodiment is a one-tube direct amplification method for detecting polymorphisms of MTHFR and / or MTRR and MTR genes.

[0078] 1. Experimental steps

[0079] 1. Materials, reagents, instruments

[0080] The detection primer probe was entrusted to the primer probe synthesis company to synthesize, and the fluorescent quantitative PCR instrument was TL988.

[0081] 2. Specimen

[0082] Saliva sample: operate according to the instructions of the saliva preservation tube

[0083] 3. Amplification of target genes

[0084] 3.1 Design of primers and probes

[0085] Design specific primers and probes based on the sequences of C677T, A1298C, A66G and A2756G. The specific primers and probes are:

[0086] SEQ ID NO.1 to SEQ ID NO.4 of the MTHFR C677T site; SEQ ID NO.5 to SEQ ID NO.8 of the MTHFR A1298C site; SEQ ID NO.9 to SEQ ID NO.12 of the MTRR A66G site; SEQ ID NO.13 to SEQ ID NO.16 of MTR A2756G site.

[0087] 3.2 Positive control: human genomic DNA.

[0088] 3.3 Neg...

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Abstract

The present invention relates to a detection composition for direct-expanded type MTHFR and / or MTRR and MTR gene polymorphisms. The detection composition comprises at least one of a detection composition 1 to a detection composition 4; wherein the detection composition 1 is sequences shown in SEQ ID NO.1 to SEQ ID NO.4 of C677T site of MTHFR; the detection composition 2 is sequences shown in SEQ ID NO.5 to SEQ ID NO.8 of A1298C site of the MTHFR; the detection composition 3 is sequences shown in SEQ ID NO.9 to SEQ ID NO.12 of A66G site of MTRR; and the detection composition 4 is sequences shown in SEQ ID NO.13 to SEQ ID NO.16 of A2756G site of MTR. The present invention also relates to a kit comprising the detection composition and a detection method. A nucleic acid extraction-free direct-expanded PCR technology is used to avoid cumbersome steps of DNA extraction, overcomes that in traditional SNP typing, one tube can only distinguish one SNP locus polymorphism, innovatively uses a multi-label TaqMan-MGB fluorescence probe, can realize that one tube can distinguish two SNP loci polymorphisms, and has good specificity and rapid sensitivity.

Description

technical field [0001] The invention belongs to the technical field of in vitro gene detection, and in particular relates to a detection composition, detection kit and detection method for direct expansion type MTHFR and / or MTRR and MTR gene polymorphism. Background technique [0002] Folic acid is a water-soluble vitamin that cannot be synthesized by the human body and must be ingested from the outside world. Although there are many foods containing folic acid, because the natural folic acid is extremely unstable and easily oxidized by irradiation and heating, the human body can not really obtain much folic acid from food. Folic acid metabolism refers to the ability of folic acid to be absorbed and utilized by the human body, mainly referring to the mutual conversion process between homocysteine ​​and methionine. In recent years, studies on the mechanism of its metabolic process have found that in addition to premature birth, miscarriage, congenital heart disease, neural t...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12Q1/6858C12N15/11
CPCC12Q1/6858C12Q1/6883C12Q2600/156C12Q2600/16C12Q2531/113C12Q2561/101C12Q2537/143
Inventor 刘荣兵徐赛涛李杨
Owner 上海酷乐生物科技有限公司
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