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Method for predicting susceptibility of coronary heart disease and kit thereof

A technology for susceptibility and coronary heart disease, applied in the fields of medical biotechnology and genetic diagnosis, can solve the problems of low sensitivity, long time, poor repeatability, etc., and achieve high sensitivity and specificity, easy operation, and stable results

Inactive Publication Date: 2011-08-17
TONGJI HOSPITAL ATTACHED TO TONGJI MEDICAL COLLEGE HUAZHONG SCI TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] At present, in the research of genetic etiology of coronary heart disease, it is generally effective to use single nucleotide polymorphism (SNP) as a genomic marker association analysis method, but the traditional PCR-enzyme digestion method of SNP analysis has a long history and is reproducible. Poor performance, low sensitivity, low accuracy, etc.

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  • Method for predicting susceptibility of coronary heart disease and kit thereof
  • Method for predicting susceptibility of coronary heart disease and kit thereof
  • Method for predicting susceptibility of coronary heart disease and kit thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0019] Example 1 Preparation of a kit for detecting susceptibility to coronary heart disease

[0020] 1. Kit components

[0021] The kit for detecting the susceptibility to coronary heart disease includes a pair of primers that can amplify the genotype of the -396D / I promoter region of the DDAH1 gene, and its Taqman probe. The components and contents are as follows, and stored at -20 degrees:

[0022] Table 1: PCR amplification reagents (200 persons)

[0023]

[0024] 2×MasterMix, supplied by ABI, has been commercialized, including the buffer components required for amplification, ABI’s gold brand enzymes, etc.

[0025] The base sequences shown in SEQ ID 2 and 3 can be seen for the sequences of F and R primers.

[0026] 2. How to use

[0027] Amplify directly on the ABI 7900 realtimePCR instrument.

[0028] Table 2: Realtime PCR reaction components (5μl reaction system)

[0029]

[0030] F primer 5'-CAGGTAAAGACCAGGAAGCCC-3' (SEQ ID 2)

[0031] R primer 5'-GGACCTCGG...

Embodiment 2

[0041] Example 2: Collection of coronary heart disease specimens and extraction of genomic DNA

[0042]The samples come from the coronary heart disease cases collected by the National 863 Project undertaken by the Institute of Hypertension of Wuhan Tongji Hospital. A total of 576 patients with coronary heart disease and 557 controls were collected. The baseline information of the population is shown in Table 3. All subjects were Han nationality, and signed informed consent, this study was also approved by the ethics committee.

[0043] According to the following method, it was used to prepare human peripheral leukocyte genomic DNA: According to the instructions of the DB-S kit produced by Japan Fuji Company, the DNA was extracted on a QG-Mini80 workstation. Genomic DNA was dissolved in TE (10mM Tris-HCl, pH 7.5; 1mM EDTA, pH 8.0)) solution, and then the mixture was quantitatively measured at 260nm absorption peak. The concentration of the DNA working solution was corrected t...

Embodiment 3

[0044] Example 3: Correlation study between DDAH1 gene-396D / I promoter region polymorphism and coronary heart disease

[0045] 1. Identification and determination of polymorphic sites

[0046] Using the above detection kits, use a 7900 PCR instrument for amplification, and finally read and analyze the amplification results.

[0047] 2. Statistical methods

[0048] SPSS 15.0 software was used for statistical data processing, and the measurement data conformed to a normal distribution and were expressed as mean ± standard deviation. The t test was used to compare the measurement data between groups. Count data, genotype and allele frequencies were compared using chi-square (χ 2 ) test calculation. The correlation between genotype and coronary heart disease was analyzed by multivariate Logistic regression to adjust the traditional risk factors of cardiovascular and cerebrovascular diseases, including gender, age, hypertension, diabetes, and hypercholesterolemia. The relative...

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Abstract

The invention provides a method for predicting the susceptibility of coronary heart disease and a kit thereof. The method comprises the following steps of: 1, picking up the genome DNA of human peripheral blood leukocytes; 2, for the polymorphic locus, namely 396D / I of the DDAH1 gene in a tested individual, which is shown as SEQ ID 1 in a sequence table, specifically amplifying the polymorphic locus from the DNA by F and R oligonucleotide primer sequences shown as the SEQ ID 2 and 3 in the sequence table, and detecting different genotypes of the DDAH1 gene, namely 396 focus by using the Taqman probe base sequence shown as SEQ ID 4 and 5 in the sequence table; and 3, introducing the I / I, D / I and D / D genotypes of the DDAH1 gene, namely 396 focus of the tested individual into a non-conditional Logistic regression analysis model to predict the susceptibility of coronary heart disease. By the method, through the specific combination of the probe and different genotypes of the polymorphic locus, the different genotypes are detected.

Description

technical field [0001] The present invention relates to a method for predicting the susceptibility of coronary heart disease (coronary artery disease, CAD), specifically by measuring human dimethylarginine dimethylamine hydrolase 1 (DDAH1) gene polymorphism site -396D / I is used to predict the susceptibility of the population to coronary heart disease. This method can be used for auxiliary diagnosis, treatment, and new drug development of diseases, and belongs to the field of medical biotechnology and genetic diagnosis. Background technique [0002] In recent years, the morbidity and mortality of coronary heart disease in my country have been rising rapidly, and it has become an important disease that has risen the fastest among the causes of death among Chinese residents and threatens the public health of our country. Traditional risk factors such as hypertension, diabetes, overweight and obesity, smoking and drinking, hyperlipidemia, etc. can cause atherosclerosis, but man...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
Inventor 汪道文丁虎吴斌
Owner TONGJI HOSPITAL ATTACHED TO TONGJI MEDICAL COLLEGE HUAZHONG SCI TECH
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