Optimization of ISSR-PCR System and Conditions for Portulaca oleracea L.

摘要

With Portulaca oleracea L.as an experimental material,its total DNA was extracted by the improved CTAB method,the ISSR-PCR primers were screened,and the ISSR-PCR reaction system and reaction conditions for P.oleracea were Optimized.The results showed that there were 8 primers suitable for IS-SR-PCR of P.oleracea.The optimal reaction system had a volume of 25μl,including 2×Taq Platinum PCR Master Mix 12.5μl,primer 2μl,ddH_2O 9.5μl,and DNA template 1μl.The optimized ISSR-PCR of P.oleracea was started with pre-denaturation at 94℃for 360 s,followed by 30 cycles of denaturation at94℃for 60 s,annealing at 54℃for 60 s and extension at 72℃ for 90 s,and completed by extension at 72℃for 300 s.