Optimization of SRAPPCR system on Poa pratensisusing orthogonal design and selection of primers

An orthogonal design was used to optimize a SRAPPCR system for Poa pratensis with 4 factors (Mg2+, dNTP, primer and Taq polymerase) at 3 levels plus the concentration of template DNA. The optimized SRAPPCR system was: 2 L 10 PCR buffer, 40 ng template DNA, Mg2+1.75 mmolL-1, dNTP 220 molL-1, primer 0.25 molL-1, Taq DNA polymerase 1.0 U in a total of 20 L reaction mixtures. With the optimized system, 43 primer combinations were selected among 100 primer combinations, which produced abundant polymorphism bands. This study optimized SRAPPCR system and selected the proper primers in P. pratensis, which would play an important role in genetic diversity analyses, map construction, germplasm identification in P. pratensis with SRAP markers.