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SNP (Single Nucleotide Polymorphism) marker for detecting diseasing risk of gestational diabetes and kit

A technology for gestational diabetes and risk of gestational diabetes, applied in the field of SNP markers and kits for detecting risk of gestational diabetes, to achieve the effect of solving early screening methods and high missed diagnosis rate, reducing prevalence rate, and lowering missed diagnosis rate

Inactive Publication Date: 2018-02-23
北京青梧桐健康科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The use of SNP to predict and assess the risk of disease, auxiliary diagnosis and prognosis assessment has been widely accepted in many fields of the medical industry, but there is no similar method for genetic risk screening of GDM.

Method used

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  • SNP (Single Nucleotide Polymorphism) marker for detecting diseasing risk of gestational diabetes and kit
  • SNP (Single Nucleotide Polymorphism) marker for detecting diseasing risk of gestational diabetes and kit
  • SNP (Single Nucleotide Polymorphism) marker for detecting diseasing risk of gestational diabetes and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] Embodiment one: DNA extraction

[0040] 1.1 DNA extraction from saliva

[0041] Saliva DNA extraction was carried out using a buccal swab genomic DNA extraction kit (brand: Kangwei Century, item number: CW0530M).

[0042] Add a certain amount of absolute ethanol to Buffer GW1 and Buffer GW2. Take 500 μl saliva sample from the sampling tube, add 300 μl Buffer GR, 20 μl Proteinase K and 300 μl Buffer GL, shake and mix, incubate with shaking at 56°C for 15 minutes, add 300 μl absolute ethanol, shake and mix; add 750 μl of the mixed solution to the collection Add 400μl Buffer GW1 to the adsorption column, centrifuge at 12000rpm for 1min, discard the solution; add 400μl Buffer GW2 to the adsorption column, centrifuge at 12000rpm for 1min, discard the solution; centrifuge at 12000rpm for 2min, and Place to dry; put the adsorption column in a new 1.5ml centrifuge tube, add 40μl Buffer GE to the suspension, let it stand for 5min, centrifuge at 12000rpm for 1min, collect the D...

Embodiment 2

[0047] Embodiment two: PCR amplification

[0048] Add the DNA of the sample to be tested extracted in 1.1 or 1.2 of Example 1 into a 384-well plate for multiplex PCR amplification. Add the reaction system (total 5 μl) according to multiplex PCR amplification into each reaction well, the reaction system:

[0049] Template DNA: 1 μl,

[0050] Primer mix: 1 μl,

[0051] 10× PCR buffer: 0.5 μl,

[0052] 25mM MgCl 2 : 0.4 μl,

[0053] 25mM dNTPs: 0.1 μl,

[0054] wxya 2 O 1.8 μL,

[0055] The balance is Hotstar Taq enzyme (5U / μl).

[0056] After the reaction system is prepared, seal it tightly with PCR parafilm to prevent the sample from evaporating, shake and mix, and centrifuge; place the sealed 384-well plate on the ABI9700 PCR instrument for the following reactions:

[0057] Pre-denaturation at 95°C for 2 minutes;

[0058] 95°C for 30s, 56°C for 30s, 72°C for 1min, 45 cycles;

[0059] 5min at 72°C, keep at 4°C;

[0060] The PCR amplification product was obtained and...

Embodiment 3

[0061] Embodiment three: SAP digestion reaction

[0062] Using the matching reagents and operating steps of the Sequenom platform, the SNP site typing results of the samples to be tested are completed. The PCR amplification product obtained in Example 2 was added to each reaction well according to the reaction system (2 μl) digested by SAP, the reaction system:

[0063] SAP×Buffer: 0.17μl,

[0064] SAP Enzyme (1U / μl): 0.3μl,

[0065] wxya 2 O: 1.53 μl;

[0066] After the reaction system is prepared, seal it tightly with PCR parafilm to prevent the sample from evaporating, shake and mix, and centrifuge; place the sealed 384-well plate on the ABI9700 PCR instrument for the following reactions:

[0067] 37°C, 40min; 85°C, 5min; 4°C hold;

[0068] The PCR product after treatment with alkaline phosphorylase was obtained and centrifuged for future use.

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Abstract

The invention discloses an SNP (Single Nucleotide Polymorphism) marker for detecting the diseasing risk of gestational diabetes mellitus. The SNP marker comprises 12 SNP sites of a personal genome, wherein the 12 SNP sites are rs7903146, rs12255372, rs 5219, rs7754840, rs4402960, rs1387153, rs10830963, rs2237892, rs1801278, rs1799884, rs1801282 and rs1063192. The SNP marker disclosed by the invention has the beneficial effects that the SNP sites selected in the invention summarize the scientific article results published at home and abroad since 1994-2016, find risk gene sites with consistentresults as detection target points, simultaneously consider the mutual action between genes and solve the problems of imperfect early screening methods and high missed-diagnosis rate for GDM on the current clinics; and after introduction of gene detection, the missed-diagnosis rate of early screening can be greatly reduced, the benefit for early intervention can be achieved and the diseasing riskof the gestational diabetes mellitus can be effectively reduced.

Description

technical field [0001] The invention relates to the technical field of gene detection, in particular to a SNP marker and a kit for detecting the risk of gestational diabetes. Background technique [0002] Due to the update of the diagnostic criteria for detecting gestational diabetes mellitus (GDM) in 2012 and the opening of the second-child policy, the prevalence of GDM in the country has reached 15-20%. GDM is a complex disease of polygenic inheritance. The heritability of the disease is over 60%, indicating that genes play a significant role in the prevalence of GDM. The risk of GDM can be predicted by the detection of susceptibility genes. [0003] The following points provide sufficient and necessary basis for the research and establishment of early screening methods for GDM: (1) The prevalence of GDM is high, and it is very harmful to pregnant women, fetuses, newborns, and postpartum women; (2) Clinically, it is not until 24 to 28 weeks that pregnant women are diagnos...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12N15/11
CPCC12Q1/6883C12Q2600/156
Inventor 牛晓强唐劼
Owner 北京青梧桐健康科技有限公司
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