摘要
以烤烟品种为材料,研究了烟草RAPD分析过程中的影响因素,包括模板浓度、Mg2+、dNTP、引物、Taq酶、循环次数、退火温度等,建立了适于烟草RAPD分析的PCR反应体系:即在25μl反应体系中,模板用量为40ng;引物浓度为0.4μM;Mg2+浓度为2.5mM;dNTP浓度为0.2mM;TaqDNA聚合酶用量为1U。扩增程序为94℃预变性5min;然后94℃变性1min,38℃复性1min,72℃延伸1.5min,39个循环;最后72℃延伸5min。
The factors influencing RAPD Analysis, including the concentration of DNA template, Mg2+, dNTP, primers, Taq polymerase, Thermal cycles and annealing temperature in tobacco were studied. An optimal PCR system for RAPD in tobacco was found: in 25μl reaction solution, contained 40ng DNA template, 0.4μM random primers, 2.5mM Mg2+, 0.2mM dNTP, 1U Taq polymerase. The amplification program was devised: 94℃ for 5min;denaturing at 94℃ for 1min; primer annealing at 38℃ for 1min, extension at 72℃ for 1.5min, 39 cycles; at last extension at 72℃ for 5min.
出处
《中国农学通报》
CSCD
2005年第5期97-100,共4页
Chinese Agricultural Science Bulletin
基金
湖南省科技厅重大科技攻关项目(01NKY1002)。
