脂联素基因突变-11377C/G检测方法建立及与2型糖尿病关系的研究

Development of an ARMS-TaqMan method to detect adiponectin-11377C/G dot mutation and correlations of-11377C/G SNP with serum adiponectin level and type 2 diabetes

目的建立脂联素基因(aPM1)-11377C/G点突变的检测方法,探讨aPM1-11377C/G点突变单核苷酸多态性(SNP)与血清脂联素(APN)水平及2型糖尿病(T2DM)的相关性。方法构建含aPM1-11377C/G点突变的野生型和突变型质粒标准品,采用等位基因特异性PCR(ARMS-PCR)方法,结合荧光探针(TaqMan Prob),通过对引物3′端正配与错配扩增效率差别的比较,建立ARMS-TaqMan检测aPM1-11377C/G点突变的方法。同时,分别对156例(男87/女69)糖尿病患者(T2DM组)和89例(男21/女68)健康对照者(NC组)的aPM1-11377C/G基因频率,以及APN、各种生化指标以及临床基本资料进行检测和调查。结果 T2DM组APN水平显著低于NC组(P<0.05)。aPM1-11377位点CC、CG、GG基因的APN水平呈趋势性降低(P<0.001)。T2DM组与对照组间的aPM1-11377位点CC、CG、GG基因频率差异有显著性(P<0.05)。T2DM组的aPM1-11377 G等位基因频率显著高于NC组(24.36%vs.19.11%,P>0.05)。结论 ARMS-TaqMan方法检测aPM1-11377C/G基因点突变具有准确、经济和适合高通量检测的特点。aPM1-11377C/G多态性与血清脂联素水平和T2DM相关。

Objective To set up a highly efficient,accurate,economical and practical screening method to detect the mutation of APN in clinical practice.Methods the ARMS-TaqMan real time PCR technique was used to detect the mutation of adiponectin.Serum adiponectin level was measured by ELISA.Results The adiponectin concentration in T2DM group is lower than that in the control group（P=0.019）.There are point mutation（C-11377G） in the region of APN gene,and it is related with the incidence of T2DM.The frequency of G allele in T2DM group is higher than that in the control group.Conclusion The aPM1 gene C-11377G and the decrease of adiponectin may be associated with T2DM.ARMS-TaqMan is a highly efficient,accurate,economical and practical screening method to detect the mutation in clinical practice.