【摘要】 目的通过对放线菌PCR扩增方法的综合改进,找到某些用常规手段难扩增的放线菌16SrDNA的最佳PCR扩增条件。方法 (1)提取放线菌基因组DNA并对其进行PCR扩增以找到最适退火温度;(2)改变PCR反应体系中模板DNA和Mg2+的浓度;(3)在反应体系中加入适量的二甲基亚砜。结果在25μL的PCR反应体系中,模板DNA浓度为10×、加入0.5μL的Mg2+和5%的二甲基亚枫、退火温度为59℃时对放线菌16SrDNA扩增效果最为理想。结论本实验研究出一种扩增放线菌16SrDNA的优化条件,为今后大批量的放线菌的分子生物学研究提供了更加高效、方便、快捷、经济的实验方法。更多还原

【Abstract】 Objective To optimize the most suitable polymerase chain reaction(PCR)amplification condition for actinomycetes strains with the adjustment and testing of different experimental methods.Methods Extracted genomic DNA of actinomycetes strains and amplify their 16SrDNA gene under various annealing temperatures.Changed the concentration of template DNA and add extra Mg2+in PCR reaction system.DMSO was added in the reaction system.Results In order to amplify 16SrDNA(about 1 500bp)of actinomycete,the most ideal condition was decided as 10×template DNA,0.5μL Mg2+and 5% DMSO,with annealing temperature of 59 ℃.Conclusion The method is efficient,convenient,fast and economical for identification and taxonomy of actinomycetes strains in large quantities.更多还原