小麦SRAP-PCR反应程序的优化及引物筛选

Program Optimization and Primer Screening of SRAP-PCR for Wheat

为建立最佳的小麦SRAP-PCR反应体系,进一步对小麦品种进行遗传多样性分析,采用L9(34)正交试验设计,对小麦SRAP-PCR程序中的变性时间、复性时间、延伸时间以及后35个循环的复性温度进行了优化试验。结果表明:当小麦SRAP-PCR扩增程序以变性30 s、复性60 s、延伸60 s,后35个循环复性温度55℃时,扩增效果最好。应用该体系从65对SRAP引物组合中筛选出扩增条带清晰、多态性丰富的23对引物组合,验证了该体系具有稳定性和可靠性,可进一步用于小麦的分子标记、辅助育种与遗传分析研究。

The purpose of this study was to determine the best system of SRAP-PCR reaction and provide a foundation for further studying the genetic diversity of wheat. Using the orthogonal design, the denaturation time, annealing time, extension time and annealing temperature after 35 cycles for wheat SRAP-PCR program were optimized. The results indicated that the best program was the denaturation time of 30 seconds,annealing time of 60 seconds,extension time of 60 seconds,and annealing temperature of 55 ℃ after 35 cycles. Under the optimum reaction conditions,23 out of 65 SRAP primer combinations were selected to verify the stability and reliability of this system. This optimized conditions of SRAP could used for further research of molecular marker,auxiliary breeding and genetic diversity in wheat.