四种小鼠肠道微生物DNA提取方法比较  

Comparison of four methods for extracting microbial DNA from mouse intestine

作　　者：吴敏娜[1] 武亚琦 屈艳[1] 魏纪东[1] 钟根深[1] 

机构地区：[1]新乡医学院基础医学院,河南新乡453003

出　　处：《生态学杂志》2015年第4期1183-1188,共6页Chinese Journal of Ecology

基　　金：国家自然科学基金项目(40901125和81201765);新乡医学院科研培育基金项目(2013ZD102)资助

摘　　要：采用异硫氰酸胍(guandine thiocyanate,GITC)法、Tiangen DNA提取试剂盒、Omega DNA提取试剂盒和广泛应用的十六烷基三甲基溴化铵(hexadecyl trimethyl ammonium bromide,CTAB)法提取小鼠粪便微生物总DNA,通过比较所提取DNA的浓度和纯度,发现粪便DNA提取试剂盒提取的DNA纯度最高但浓度最低;CTAB法所得的DNA浓度最高但纯度最低;GITC法所得DNA的浓度高于粪便DNA提取试剂盒,纯度高于CTAB法。通过变性梯度凝胶电泳(16S r DNA-PCR-DGGE)指纹图谱分析技术进一步比较了各种提取方法所代表微生物群落的丰富度和多样性。结果表明,GITC法提取得到的DNA所代表细菌的丰富度和多样性显著高于其他3种方法。本实验所建立的GITC法可更全面地反映肠道微生物的多样性和群落结构,是一种较为理想的粪便微生物DNA提取方法。The total microbial DNA of mice feces was extracted by GITC（ guandine thiocyanate）method,CTAB（ hexadecyl trimethyl ammonium bromide） method,Tiangen and Omega fecal DNA extraction kits. The yield and purity of the extracted DNA were determined by spectrophotometer of Nano Drop 2000. The results showed that the DNA extracted by the commercial DNA extraction kits was of the highest purity but of lowest yield,while the DNA with highest yield and lowest purity was obtained by CTAB method. The DNA extracted from mice feces by using GITC method showed higher purity than that retrieved from CTAB method,and higher yield than that extracted by commercial DNA extraction kits. The bacterial richness and diversity were further compared among the four methods by using 16 S r DNA-PCR-DGGE fingerprinting method. Results revealed that significantly higher bacterial richness and diversity were observed in the community extracted from GITC method than that from the other three methods. In conclusion,the GITC method developed in our lab had some advantages as a fecal DNA extraction protocol,with higher DNA quality and quantity,specifically,higher richness and diversity of intestinal microbiota,compared with CTAB method,Tiangen and Omega fecal DNA extraction kits.

关 键 词：DNA提取 肠道微生物 粪便 变性梯度凝胶电泳(DGGE) 

分 类 号：Q933[生物学—微生物学]