苹果自交不亲和基因PCR扩增程序优化

Optimized Study on PCR Amplification Procedure of the Apple Self-incompatibility Gene

通过研究PCR反应体系(DNA、Primer、Mg2+、dNTP、Taq聚合酶、buffer)及退火温度对苹果自交不亲和基因(S基因)扩增结果影响的分析,构建了适合于苹果自交不亲和基因的PCR扩增程序的优化体系。结果表明:在20μL的反应体系中,模板DNA的最适含量为40ng,dNTP最适浓度为0.25mmol/L,而Primer最适浓度为0.4μmol/L,Mg2+的最适浓度为2.5mmol/L,Taq聚合酶最适浓度为1U,buffer最适浓度为1×buffer,最适退火温度为50℃。

Through analyzing the effect of PCR system （DNA, Primer, Mg^2＋, dNTP, Taq polymerase, and buffer） and annealing temperature on amplification of apple self-incompatibility gene （s-gene）, we constructed a optimized system of PCR amplification procedure which was fit for apple self-incompatibility gene. The result showed that the optimized content in 20 μL system contained 40 ng DNA template, 0.25 mmol/L dNTP, 0.4 μmol/L primer, 2.5 mmol/L Mg^2＋, 1 U Taq polymerase, lxbuffer, and the optimized annealing temperature was 50 ℃.