The Optimization of the SSR Reaction System for Wild European Plum (Prunus domestica L.)

摘要

The research aims at obtaining the optimal SSR-PCR system for Wild European Plum,which is a kind of endangered fruit trees.The Wild European Plum was used as the research object,and an experimental design was employed to evaluate five factors (template DNA,dNTP,Mg2+,Taq DNA polymerase and primer) at six different levels.The results indicate that the optimal SSR-PCR reaction system for Wild European Plum was determined as follows:the optimal system of terminal volume 15μ1 consisted of 0.4U Taq DNA polymerase,0.4mM primers,30-50ng tem-plate DNA,0.3mM dNTP and 2.0mM Mg2+.The suitable thermal cycling conditions are melting at 94°C for 5 min initially,followed by 35 cycles at 94°Cfor 30 s,50-59°Cfor 45 s and 72°Cfor 60 s,then keep the reaction mixture at 4°C after a final extension step of 72°C for 7-10 min.The optimized system would be effective as a solid foundation for Wild European Plum SSR analysis.