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中国农学通报 ›› 2005, Vol. 21 ›› Issue (5): 97-97.

所属专题: 烟草种植与生产

• 目次 • 上一篇    下一篇

烟草RAPD反应体系的建立与优化研究

杨友才,周清明,尹晗琪   

  • 出版日期:2005-05-05 发布日期:2005-05-05

Establishment and Optimization of RAPD Reaction System in Tobacco

Yang Youcai, Zhou Qingmin, Yin Hanqi   

  • Online:2005-05-05 Published:2005-05-05

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;Taq DNA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。

Abstract: 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.