PCR-RFLP技术检测SUMO4基因163A/G多态性的实验条件研究  

The Experimental Conditions for SUMO4 Gene 163A/G Polymorphism Detection with PCR-RFLP

作　　者：李斌[1] 李会芳[1] 王玉明[2] 周晰溪[1] 张娴[1] 宋滇平[1] 

机构地区：[1]昆明医学院第一附属医院糖尿病科,云南昆明650032 [2]昆明医学院第一附属医院检验科,云南昆明650032

出　　处：《昆明医学院学报》2010年第5期4-7,共4页Journal of Kunming Medical College

基　　金：国家自然科学基金资助资助(30760087)

摘　　要：目的确定PCR-RFLP技术检测小泛素样修饰蛋白4(SUMO4)163A/G多态性的最适实验条件.方法对影响SUMO4基因PCR反应的主要因素设定系列浓度梯度分别进行研究.结果最适变性温度为94℃;最适退火温度为55℃;最适引物浓度为0.25μmol/L;最适Mg2+浓度为1.5mmol/L;最适dNTP浓度为0.25mmol/L;以2.5U为最适Taq酶量;酶切体系为31μL体系中加10μL产物用10U的酶消化.结论最佳实验条件的摸索是进行糖尿病实验研究的关键.Objective To determine the optimum expermental conditions for the polymorphism of SUMO4 gene 163A/G detected by polymerase chain reaction-restriction fragment length polymorphism（PCR-RFLP） . Methods The main influential factors were detected by PCR in serial concentration gradient and electrophoresis,the main factors were detected by RFLP in different system.Results The optimum conditions：denaturation temperature was 94 ℃;annealing temperature was 55 ℃;primer concentration was 0.25 umol/L;magnesium concentration was 1.5 mmol/L;concentration of dNTP was 0.25 mmol/L;Taq enzyme was 2.5 U;the effective system was 31 uL including 10 uL DNA solution and 10 U enzyme.Conclution The key way to a large empirical study is to detect the optimal experiment conditions of diabetes.

关 键 词：聚合酶链反应 限制性片段长度多态性 小泛素样修饰蛋白4 实验条件 

分 类 号：R365[医药卫生—病理学]