Abstract:

Objective To assess the clinical application of polymerase chain reaction（PCR） melting curve method in determining hypertensive drug-related gene mutations. Methods Totally,100 hypertension patients were enrolled,and the single nucleotide polymorphism（SNP） sites of hypertensive drug-related gene mutations,including cytochrome P450 2D6（CYP2D6）rs1065852,cytochrome P450 2C9（CYP2C9）rs1057910,betal-adrenergic receptor-1（ADRB1）rs1801253,angiotensin Ⅱ receptor 1（AGTR1）rs5186 and angiotensin-converting enzyme（ACE）rs1799752,were determined. DNA sequencing was performed. The accuracy,repeatability,anti-interference and the genotype frequency of PCR melting curve method were evaluated. Results The consistency of PCR melting curve method and DNA sequencing was 100%. The extremum values（R） of the Tm about 5 gene sites were all <0.5 ℃. Hemoglobin（Hb）≤2 g/L,bilirubin（Bil）≤342 μmol/L,triglyceride（TG）≤37 mmol/L had no effect on PCR melting curve method. The coefficient of variation（CV）of Tm was ≤5%. In 100 hypertension patients,there was 1 case of AGTR1（c.1166A>C） and 1 case of CYP2C9*3（c.1075A>C）homozygous mutations. The genotype frequencies of CYP2D6*10 and CYP2C9*3 were 39.5% and 5.5%,respectively. The genotype frequencies of C allele of mutant ADRB1,C allele of mutant AGTR1 and D allele of ACE deletion were 62.0%,5.0% and 35.0%,respectively. Conclusions PCR melting curve method is simple and has low requirements for equipments,and it can be used as a routine method for determining anti-hypertensive drug-related gene mutations.

Key words: Polymerase chain reaction melting curve method, Hypertensive drug-related gene, Single nucleotide polymorphism