Optimization for SRAP—PCR system in Cynodon dactylon and selection of primers
Cynodon dactylon genomic DNA from leaf tissue was isolated according to the SDS proceduce. An orthogonal diagram as experimental design was applied to study the effects of three levels of Mg2+, dNTP, primers and Taq DNA polymerase respectively on the reaction system of SRAP, and the SRAP reaction system was finally optimized by analyzed effects of the concentrations of DNA template. The results showed that all factors had the significant effect on the result of SRAP-PCR. And a better amplification of SRAP-PCR was obtained with the reaction system for C. dactylon containing 2 μL 10×buffer, 40 ng genomic templates DNA, 1.25 mmol/L Mg2+ , 260 μmol/L dNTP, 0. 2 μmol/L Primer, 1.0 U Taq DNA polymerase in the total volume of 20 μL. At the same time, 34 primer combinations were selected with the optimized system among 90 primer combinations, which had abundant polymorphism bands. This study would be conducted to the optimized SRAP-PCR system and selection of primers which would play an important role in C. dactylon genetic diversity analysis, map construction, gene localization of important traits, germplasm identification in C. dactylon with SRAP markers.
Cynodon dactylonSRAP markerorthongonal designoptimization of systemprimers
国家科技期刊平台
NSTL
万方数据
维普
