[Objective] The aim was to investigate the optimal conditions of SRAP molecular marker used in the analysis on Fagopyrum tataricum（L.）Gaertn.[Method]SRAP-PCR amplification system on Fagopyrum tataricum was optimized...[Objective] The aim was to investigate the optimal conditions of SRAP molecular marker used in the analysis on Fagopyrum tataricum（L.）Gaertn.[Method]SRAP-PCR amplification system on Fagopyrum tataricum was optimized by interactive orthogonal design L27（313）in 5 elements（Mg2＋,dNTP,Taq DNA polymerase,template DNA and primer）at 3 levels.And the non-denaturing and denaturing PAGE detection methods were compared.The comparative test of DYCZ-24F and DYCZ-20C electrophoresis operating systems was carried out.[Result]The effects of four single-factor（Mg2＋,dNTP,Taq DNA polymerase and primer）and two interactions（Mg2＋×dNTP,Mg2＋×Taq DNA polymerase）on tartary buckwheat SRAP-PCR were significant.An optimal reaction system was established containing 1.5 mmol/L Mg2＋,0.2 mmol/L dNTP,1.5 u Taq DNA polymerase,40 ng DNA,0.25 μmol/L primer and 2 μl 10×buffer.Seven samples of tartary buckwheat were amplified using this system,and electrophoresis results showed clear bands,high level of polymorphism and good reproducibility.The PCR products were tested by denaturing and non-denaturing PAGE,and the results showed that the non-denaturing PAGE,DYCZ-24F operating system was more suitable for SRAP analysis.[Conclusion]This study established a foundation for the construction of SRAP genetic map of tartary buckwheat.展开更多
基金Supported by National Natural Science Foundation of China(30771310)~~
文摘[Objective] The aim was to investigate the optimal conditions of SRAP molecular marker used in the analysis on Fagopyrum tataricum（L.）Gaertn.[Method]SRAP-PCR amplification system on Fagopyrum tataricum was optimized by interactive orthogonal design L27（313）in 5 elements（Mg2＋,dNTP,Taq DNA polymerase,template DNA and primer）at 3 levels.And the non-denaturing and denaturing PAGE detection methods were compared.The comparative test of DYCZ-24F and DYCZ-20C electrophoresis operating systems was carried out.[Result]The effects of four single-factor（Mg2＋,dNTP,Taq DNA polymerase and primer）and two interactions（Mg2＋×dNTP,Mg2＋×Taq DNA polymerase）on tartary buckwheat SRAP-PCR were significant.An optimal reaction system was established containing 1.5 mmol/L Mg2＋,0.2 mmol/L dNTP,1.5 u Taq DNA polymerase,40 ng DNA,0.25 μmol/L primer and 2 μl 10×buffer.Seven samples of tartary buckwheat were amplified using this system,and electrophoresis results showed clear bands,high level of polymorphism and good reproducibility.The PCR products were tested by denaturing and non-denaturing PAGE,and the results showed that the non-denaturing PAGE,DYCZ-24F operating system was more suitable for SRAP analysis.[Conclusion]This study established a foundation for the construction of SRAP genetic map of tartary buckwheat.
