沙棘SCoT-PCR反应体系的优化及引物筛选

Optimization of SCoT PCR Reaction System and Selection of Primers for Hippophae rhamnoides

为获得沙棘SCoT-PCR最佳反应体系并筛选出适用引物,本研究采用正交试验设计与单因素试验设计的方法对其反应体系进行优化,并在此基础上对设计的80条引物进行筛选。试验结果表明:沙棘SCoT-PCR最佳反应体系(20μL)为:2×Taq PCR预混试剂Ⅱ用量9.8μL,模板DNA用量25 ng,引物浓度0.9μmol/L;从设计的80条SCoT引物中筛选出24条扩增条带清晰、多态性较高的引物。本研究结果可为沙棘遗传多样性、遗传图谱构建、品种指纹图谱的构建等研究提供相关依据。

In order to obtain the best SCoT-PCR reaction system of Hippophae rhamnoides and select suitable primers,combined orthogonal design and single factor design were used to optimize the reaction systemand on this basis,the 80 primers designed were screened.The results showed that the optimal reaction system(20μL)of SCoTPCR for H.rhamnoides was as follows:9.8μL 2×Taq PCR MasterMixⅡ,25 ng template DNA,and 0.9μmol/L primer;24 primers with clear amplification bands and high polymorphism were screened out from 80 primers.It can provide relevant basis for the genetic diversity,genetic map construction and variety fingerprint construction of H.rhamnoides.