Optimization of SSR-PCR Reaction System in Jatropha curcas

Key words: 
• Jatropha curcas /
• DNA extraction /
• SSR-PCR reaction /
• system optimization

Abstract: Genomic DNA of Jatropha curcas was extracted by improved CTAB method from J. curcas leaves,and detected through agarose gel electrophoresis and SSR-PCR amplification. The results showed that genomic DNA of J. curcas extracted by the modified CTAB method had good integrity,high purity,and hence could be used as a template of SSR markers. At the same time,4 factors influencing PCR amplification: d NTPs,Taq DNA polymerase,primers and Mg²⁺concentration were optimized to obtain a reaction system most suitable for SSR-PCR amplification and SSR-PCR amplification procedure for J. curcas. In this optimal reaction system with a total volume of 20μL the template DNA was 100 ng,d NTPs 0. 2 mmol·L^-1,Taq DNA polymerase 100 U·m L^-1,primer 0. 6μmol·L^-1 and Mg²⁺1. 5 mmol·L^-1,and the suitable amplification program included 94℃ pre degeneration for 3min at 94℃,denaturation for 1 min at 94℃,annealing for 1 min at 51- 53℃（ annealing temperature changed with primers）,extension for 2 min at 72℃,35 cycles,extension for 10 min at 72℃.