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Kit for rapid detection of SNP loci associated with hypertension treatment and prognosis

A kit, high blood pressure technology, applied in the direction of recombinant DNA technology, microbial measurement/testing, DNA/RNA fragments, etc., can solve the high-level problems of non-fatal myocardial infarction, achieve short time-consuming, simple operation, and broad application prospects Effect

Active Publication Date: 2019-08-16
JIANGSU VOCATIONAL COLLEGE OF MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, those patients with the Gln / Gln(G / G) genotype were at significantly higher risk of all-cause death, non-fatal myocardial infarction, and non-fatal stroke when treated with verapamil extended-release Patients treated with Puli

Method used

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  • Kit for rapid detection of SNP loci associated with hypertension treatment and prognosis
  • Kit for rapid detection of SNP loci associated with hypertension treatment and prognosis
  • Kit for rapid detection of SNP loci associated with hypertension treatment and prognosis

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] The working effect detection of embodiment 1-design primer and the elimination of false positive

[0054] 1. Prepare the positive standard plasmid KFSA-native containing the sequence shown in SEQ ID NO: 1 and the allelic standard plasmid KFSA-allele containing the sequence shown in SEQ ID NO: 2 by using a small plasmid extraction kit, using ultra-trace protein nucleic acid The analyzer detects its concentration and purity, records the original data, and stores the DNA whose concentration and purity meet the requirements in a -20°C refrigerator for future use.

[0055] 2. The working efficiency of the designed primers for rs4253238 of KLKB1 gene, rs2731672 of F12 gene, rs16982743 of SIGLEC12 gene and rs2004776 of AGT gene were detected by conventional PCR method. The reaction system is:

[0056]

[0057] The reaction conditions were as follows: pre-denaturation at 94°C for 10 min, denaturation at 94°C for 30 s, annealing at 60.5°C for 20 s, extension at 72°C for 32 s...

Embodiment 2

[0061] Embodiment 2-standard reaction system and conditions of the kit of the present invention, and the establishment of the basis for judging the results

[0062] 1. Preparation of positive standard plasmid KFSA-native

[0063] Using the plasmid small extraction kit, the operation process was carried out according to the kit instructions, and the positive standard plasmid KFSA-native containing the sequence of SEQ ID NO: 1 was prepared, and its concentration and purity were detected by an ultra-micro protein nucleic acid analyzer, and the original data was recorded. DNA whose concentration and purity met the requirements was stored in a -20°C refrigerator for later use.

[0064] 2. Real-time quantitative PCR amplification of the target gene

[0065] Use primers for the rs4253238 site of the KLKB1 gene, primers for the rs2731672 site of the F12 gene, primers for the rs16982743 site of the SIGLEC12 gene, and primers for the rs2004776 site of the AGT gene in a molar ratio of 2...

Embodiment 3

[0070] Example 3 - Using the kit of the present invention to analyze SNP sites in patients with hypertension

[0071] 1. Collection of samples and extraction of whole genome DNA

[0072] Randomly collect 3 mL of peripheral blood from hypertensive patients with EDTA anticoagulant tubes, and use Vazyme’s FastPureBlood DNA Isolation Mini Kit to extract the whole genome DNA of the subject. The nucleic acid analyzer detects its concentration and purity, records the original data, and stores the DNA whose concentration and purity meet the requirements in a -20°C refrigerator for future use.

[0073] 2. Real-time quantitative PCR amplification of the target gene

[0074] Use primers for the rs4253238 site of the KLKB1 gene, primers for the rs2731672 site of the F12 gene, primers for the rs16982743 site of the SIGLEC12 gene, and primers for the rs2004776 site of the AGT gene in a molar ratio of 20-22:1-1.5:1-1.5:1. The primer mixture was obtained for real-time quantitative PCR ampli...

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Abstract

The invention discloses a kit for rapid detection of SNP loci associated with hypertension treatment and prognosis. The kit comprises four pairs of primers for detecting four SNP polymorphism sites corresponding to the KLKB1 / F12 / SIGLEC12 / AGT genes. The kit of the present invention is based on the nsSNP genotyping Multi-AS-PCR primer design, through real-time fluorescent quantitative PCR amplification, according to the characteristic peak of a melting curve, the SNP typing of hypertension treatment and prognosis risk gene is provided, which provides a basis for personalized diagnosis and treatment of hypertensive patients. The kit of the invention has the characteristics of simple operation, low cost, short time consumption and reliable result, and has broad application prospects.

Description

technical field [0001] The invention belongs to the field of molecular biology detection and the field of cardiovascular and cerebrovascular diseases, and in particular relates to a kit for rapidly detecting SNP sites related to hypertension treatment and prognosis. Background technique [0002] In 2014, it was estimated that there were 290 million cardiovascular disease patients nationwide, including 270 million hypertensive patients, at least 7 million stroke patients, and 2.5 million myocardial infarction patients. Through the research on the occurrence and development mechanism of cardiovascular disease complications, it is found that genetic factors play an important role. The analysis of susceptible, prevalent and clinical cases with different symptoms found that single nucleotide polymorphisms (SNPs) are closely related to the occurrence and development of diseases, especially non-synonymous SNPs (nsSNPs). [0003] Nejatizadeh et al. (2008) found a common SNP (G-6A, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6883C12Q1/6858C12N15/11
CPCC12Q1/6883C12Q1/6858C12Q2600/156C12Q2600/118C12Q2561/113C12Q2545/114C12Q2531/113C12Q2563/107C12Q2527/107
Inventor 钱炳俊俞黎黎赵霄纪阳丁子璇岳杨范晨
Owner JIANGSU VOCATIONAL COLLEGE OF MEDICINE