Establishment Of ACE Genotyping Method Based On PCR-goldmag Lateral Assay And The Applications In Detection Of Clinical Sample

Background : Angiotensin converting enzyme(ACE)gene,as a candidate gene for hypertension and cardiovascular and cerebrovascular diseases,has received extensive attention in recent years.Angiotensin converting enzyme(ACE)is an important component of the renin angiotensin system.Studies have shown that ACE gene insertion/deletion polymorphisms are closely related to the efficacy of medication and the genetic susceptibility of cardiovascular and cerebrovascular diseases in hypertensive patients.Therefore,the detection of ACE gene polymorphism can guide the use of drugs in patients with hypertension with different genotypes,and can also predict and prevent cardiovascular and cerebrovascular diseases.Purpose: Based on the gold magnetic particle chromatography system,a rapid genotyping method for multi-sample ACE gene was established.The systematic performance of the method was evaluated,and the accuracy and applicability of the method were evaluated by combining with sequencing and agarose gel electrophoresis.In order to use this method to detect ACE genotypes in patients with hypertension and to guide drug use.At the same time,this method was used to analyze the association between coronary heart disease and stroke clinical samples and the locus of the gene,and to explore the association between the insertion / deletion polymorphism of ACE gene and the two genes.Methods and main results:1.Specific primers were designed for the insertion of ACE gene deletion fragments.The primers were screened and marked,and the DNA typing method of ACE gene was established.The method of sequencing and agarose gel electrophoresis was used to verify the detection system,and the performance of the method was evaluated.The detection method is simple,rapid,and sensitive.After a simple PCR amplification,chromatographic analysis can be performed visually for 2 to 5 minutes.2.Based on the establishment of DNA typing method for ACE gene,using whole blood as template,the whole blood direct spread sequence detection method of ACE gene was established,and the detection system was optimized.The clinical samples were compared by sequencing and agarose gel electrophoresis typing,and the performance evaluation of the methodology was completed.The establishment of whole blood direct spreading system of ACE gene saves the tedious steps of nucleic acid extraction and avoids the risk of cross-contamination in the process of nucleic acid sample extraction.The whole detection process is about 1 hour and 20 min,which has great clinical application value.3.Healthy clinical samples were used as control group,coronary heart disease(CAD)and ischemic stroke(ISI)clinical samples as case groups.Genotypes were detected by the established ACE genotyping method,and the results were statistically analyzed.The results showed that there was no significant difference in allele and genotype frequency between control group and CAD group and is.The reason may be that coronary heart disease and stroke is a complex multi gene disease.Multiple joint analysis of multiple related loci is needed to provide more detailed information,so as to provide a clinical reference for the study of ACE gene polymorphism and genetic susceptibility between stroke and coronary heart disease.