平菇基因组DNA提取方法的研究  

Study on the methods of extraction of Pleurotus ostreatus genomic DNA

作　　者：闫燕[1,2,3] 许文涛[2,3] 苏春元[3] 罗云波[3] 王一南[3] 谷新晰[1] 戴蕴青[2,3] 田洪涛[1] 

机构地区：[1]河北农业大学食品科技学院,河北保定071001 [2]中国农业大学食品科学与营养工程学院,北京100083 [3]农业部转基因生物食用安全监督检验测试中心(北京),北京100083

出　　处：《食品工业科技》2011年第9期190-193,共4页Science and Technology of Food Industry

基　　金：国家高技术研究发展计划(863)项目(2006AA10Z440);国家自然基金(30800770);转基因生物重大专项(2008ZX08012-001)

摘　　要：以实验室自行培育的平菇为实验材料,分别使用CTAB法、SDS-CTAB法以及高盐酶解法提取平菇基因组DNA,并通过紫外分光光度计、限制性酶切及PCR检测来衡量基因组质量。结果表明,使用CTAB法很难得到高质量的基因组DNA,OD260/280仅为1.5左右,而且伴有较多的杂质,电泳条带拖尾严重;使用EcoRI限制性酶对其进行酶切分析,效率较低,同时PCR检测不能得到有效扩增;而使用优化后的SDS-CTAB法及高盐酶解法提取平菇DNA,能够获得质量高、纯度好的基因组DNA,OD260/280基本保持在1.8～2.0之间,电泳条带较为清晰均一,使用EcoRI限制性酶对其进行酶切分析,酶切效果较好,并且DNA质量能够满足PCR检测的模板要求。说明SDS-CTAB法以及高盐酶解法提取的平菇DNA能够满足分子水平操作的要求,两者相比,前者成本较低,后者产率较高。Pleurotus ostreatus（culture by ourselves）genomic DNA were extracted by three methods,which were the methods of CTAB method,SDS-CTAB method,and high-concentration salt precipitation method,respectively,analyzed by ultraviolet spectrophotometer,digested by restriction enzymes of EcoRI and PCR amplified. The results proved Pleurotus ostreatus genomic DNA was really difficult to isolated genomic DNA by CTAB method,the ratio of OD260/280 was only 1. 5,digested by EcoRI inefficiency and PCR amplification useless. The other two methods were proved efficiently,the ratio of OD260/280 was between 1. 8～2. 0 and had a clear fragment through agarose gel electrophoresis,digested by EcoRI efficiency and could be used as template for PCR amplification analysis,while the effect of extract genomic DNA by SDS-CTAB purification method and high-concentration salt enzymolysis method could be used for downstream analyses,the former was lower cost and the latter was more efficiency.

关 键 词：平菇 基因组DNA 提取方法 

分 类 号：TS255.1[轻工技术与工程—农产品加工及贮藏工程]