4种不同土壤微生物DNA提取方法对DGGE分析微生物群落的影响

摘要

The Zhou, Martin, Bourrain and Kit methods were used to extract microbial DNA in paddy soil. They were evaluated based on the amplification of bacterial 16S rRNA Va variable region amplified by the primer pair GC338f-530R and the fungal 18S rRNA region amplified by the primer pair NS1-GCFung and the subsequent DGGE analysis of amplified products. The results showed that the Martin and the Kit methods satisfied the analytical requirements of soil microbial diversity. The Kit method was simpler and faster, and the quality of its extracted DNA higher, but the yield and purity of DNA extracted were lower and the cost greater than the Martin method. However, the deficiencies did not appear to impose significant negative effect on the PCR amplification and DGGE analysis that followed.%分别应用高盐法、玻璃珠破碎法、冻融法和试剂盒法4种不同土壤微生物DNA提取方法提取水稻稻田土壤微生物DNA,并通过细菌16S rRNA V3区通用引物GC338f-530R和真菌18S rRNA特异性引物NSl-GCFung进行PCR扩增结合变性梯度凝胶电泳(CGGE)分析,对4种DNA提取方法进行评价.结果表明,玻璃珠破碎法和试剂盒法提取的DNA均能满足土壤微生物多样性分析的要求;其中试剂盒方法操作简单,提取的DNA质量较高,但DNA产量较低且成本昂贵；玻璃珠破碎法用时较长,步骤繁琐,DNA产量较高,纯度较低,但对后续PCR扩增和DGGE分析没有明显影响,且成本低廉.