一种基于PCR分析多样性的小鼠肠道微生物宏基因组提取方法

目的建立一种基于PCR分析分子多样性的小鼠肠道菌群宏基因组提取方法。方法比较、综合国内外小鼠肠道菌群宏基因组的提取方法后建立一种新方法，小鼠肠道内容物经丙酮洗涤，差速离心，溶菌酶、SDS裂解，CTAB处理，酚／氯仿抽提后可得到高质量的DNA，通过紫外分光光度计、琼脂糖凝胶电泳、细菌通用引物PCR和扩增核糖体限制性酶切片段分析（ARDRA）等检测该方法的实用性。结果该方法获得的小鼠肠道菌群宏基因组DNA大小在23kb左右，A260／A280在1．8—2．0，经细菌通用引物PCR后能得到适用于ARDRA的目的产物。结论该方法经济适用性较强，具备一定的应用价值。

A metagenome DNA extracting method of intestinal flora in mice for molecular diversity analysis based on PCR technology

Objective To establish a method of metagenome DNA extraction of intestinal flora in mice for molecular diversity analysis based on PCR.Method The faecal samples were collected from intestinal tracts of mice.These samples were washed by acetone,and then centrifuged in different speeds.The cells were lysed by lysozyme and SDS,treated by CTAB,and then extracted with phenol/chloroform.The metagenome DNA of bacterial species was determined by a UV spectrophotometer,agarose gel electrophoresis.The 16S rDNA was amplified from these DNA samples through a set of bacterial universal primers.The PCR products were analyzed by Amplifed Ribosomal DNA Restriction Analysis(ARDRA).Result The metagenome DNA of intestinal flora in mice extracted by this method was about 23 kb;the A260/A280 of DNA was between 1.8 and 2.0;the PCR products were about 1.4 kb and can be applied to ARDRA for further analysis.Conclusion This method is cost-effective and easy,which is suitable for metagenome DNA extraction of intestinal flora.