竹黄菌基因组RAPD分子标记体系的建立

Establishment of RAPD Molecular Marker Reaction System for Shiraia bambusicola

目的:建立随机引物扩增多态性DNA(RAPD)分子标记体系,为研究华东地区不同地理居群竹黄菌Shiraia bambusicola的遗传多样性奠定基础。方法:应用SDS法提取竹黄菌基因组DNA,并优化影响RAPD反应的条件因素。结果:竹黄RAPD的最佳反应体系为:25μlPCR反应体积,10×PCR缓冲液2.5μl,1.0UTaq酶,50ng模板DNA,2mmol/LMg2+,0.2mmol/LdNTPs,0.4μmol/L随机引物。PCR循环程序为:94℃预变性4min,然后,94℃变性1min,38℃退火70s,72℃延伸90s,40次循环,最后72℃延伸6min,12℃保存。结论:建立了竹黄菌Shiraia bambusicola的最佳RAPD分子标记体系,可获得良好的竹黄菌RAPD指纹图谱。

Objective：Random amplified polymorphic DNA （ RAPD） molecular marker reaction system was established to analyze the genetic diversity of different geographic populations Shiraia bambusicola from East China.Method：The genomic DNA was extracted using the SDS method and then the optimization of its RAPD reaction system was studied.Result：The optimal PCR system for RAPD analysis was as follows：2.5μl 10×PCR buffer,1.0 U Taq polymerase,50ng DNA template,2mmol/L Mg2 ＋ ,0.2mmol/L dNTPs,0.4μmol/L random primer,in 25μl reaction system.PCR program was 4 min.94℃ for predenaturation,then followed by 40 cycles,each with 1 min.at 94℃ for denaturation,70s at 38℃for annealing,90s at 72℃ for extension,finally extension at 72℃ for 6 min.Conclusion：It provided a reliable method of the best S.bambusicola RAPD reaction system for obtaining PAPD fingerprint of S.bambusicola