PCR-RLFP方法检测IL-6基因启动子区-634C/G多态性的实验条件研究

Study of the Trial Conditions of PCR-RLFP Detecting Polymorphism of IL-6 Gene Promoter Region-634C/G

目的探讨PCR-RFLP技术判断IL-6基因启动子区-634C/G多态性的最适实验条件.方法对影响PCR的部分因素和影响RFLP方法的一些因素进行研究.结果PCR扩增IL-6基因启动子区的最适条件为:引物浓度为0.2μmol/L;Taq酶量为1.5 U;dNTP浓度为120μmol/L;Mg2+浓度为2.0 mmol/L.最经济有效的酶切体系为20μL体系中加4μL产物用2.5 U的酶消化9 h以上.结论最佳实验条件的探索是进行大批量实验研究的关键.

Objective To explore the most optimum trial conditions of polymerase chain reaction - restriction fragment length polymorphism to detect the polymorphism of IL - 6 gene promoter region - 634C/G. Methods To study the factors influencing polymerase chain reaction - restriction fragment length polymorphism. Resuits The optimum conditions of PCR method which amplified IL - 6 gene promoter region were each of primers concentration of 0. 2μmol/L, Taq polymerase amount of 1.5 U, dNTP concentration of 120μmol/L and Mg2＋ concentration of 2. 0 mmol/L . The most economic and effective system was 20μL including 4 μL DNA solution and 2. 5 U enzyme. Enzyme digestion time was more than 9 hours. Conclusion The key way is to detect the optinal experiment conditions in a large empirical study.