Abstract:

A SSR marker detection system for sesame (Sesamum indicum L.) by optimizing the amplification reaction components, cycling parameters and the electrophoresis detection was established. DNA samples from five sesame cultivars were used as template for this study. The optimized PCR system contained the following components in a volume of 10 μL: 25 mmol/L Mg2+ 1.2 μL, 10 mmol/L dNTPs 0.3 μL, 5 U/μL Taq polymerase 0.3 μL, 50 ng/μL Primer 0.9 μL, 10×PCR reaction buffer 0.7 μL, 25 ng/μL DNA template 2.5 μL, ddH2O 4.1 μL. PCR amplification was conducted in three phases. Firstly, DNA was denatured for 3 min at 94℃ and then 10 cycles of 94℃ 1 min, 60℃ 30 s and 72℃ 45 s. Secondly, another 30 cycles of 94℃ 30 s, 57℃ 30 s, 72℃ 45 s were performed. Finally, the amplification was ended in 5 min at 72℃. The 6% denaturing polyacrylamide gel electrophoresis and silver staining method were employed for the detecting of banding profile. With the above detection system, 35 SSR markers were screened across 5 sesame cultivars and found that 24 markers could reveal two or more polymorphic bands. The usefulness of the optimized detection system and the set of SSR markers for cultivar identification were verified in the analysis of other 20 sesame cultivars.