油页岩微生物总DNA的提取方法  

Extraction Methods of Microbial Total DNA from Oil Shales

作　　者：蒋绍妍 王文星[1,2] 薛向欣[1] 侯晓娇 

机构地区：[1]东北大学材料与冶金学院,辽宁沈阳110819 [2]东北大学生命科学与健康学院,辽宁沈阳110819

出　　处：《东北大学学报（自然科学版）》2014年第4期588-592,共5页Journal of Northeastern University(Natural Science)

基　　金：国家自然科学基金资助项目(51204055);中国博士后科学基金资助项目(20100481205)

摘　　要：为建立油页岩微生物总DNA提取方法，以两矿区的6个样品为材料，采用SDS高盐、SDS液氮研磨、SDS反复冻融、SDS异硫氰酸胍和试剂盒提取法分别对总DNA进行提取．结果表明，各方法提取效果差异很大，改进的SDS高盐提取法效果最好，各样品均得到约23kb较完整片段，且产率显著高于其他方法，达91．44～386．85ng·g^-1干样；以未纯化的DNA为模板，对16SrDNA进行PCR—DGGE，得到了相应的扩增产物及DGGE指纹图谱，说明该法适合油页岩样品．SDS液氮研磨和SDS反复冻融提取法仅能得到部分样品总DNA，且产率和纯度较低，而SDS异硫氰酸胍和试剂盒提取法无DNA提出，不适合此类样品．In order to develop an appropriate method for extracting microbial total DNA from oil shales, 5 methods （SDS-high-salt extraction, SDS-grinding-in-liquid-nitrogen extraction, SDS repeated freeze-thaw extraction, SDS guanidinium isothiocyanate extraction and kit extraction ） were tested to extract total DNA of 6 different samples from two mines, respectively. The results showed that the total DNA extraction effects of five methods vary significantly. The improved SDS-high-salt extraction method is the best, which can obtain total DNA with about 23 kb length and 91.44 -386. 85 ng-g- 1 dry sample. The yield rate is significantly higher than other methods. Then, the total DNAs without purification process were used as amplification templates, and 16S rDNA was carried out PCR amplification and DGGE electrophoresis, the corresponding PCR products and DGGE fingerprint were obtained. This shows that the improved SDS-high-salt extraction method is suitable for oil shale samples. SDS-grinding-in-liquid-nitrogen extraction and SDS repeated freeze-thaw extraction are only able to extract microbial total DNA of part of the samples, and with low yield and purity. SDS guanidinium isothiocyanate extraction and kit extraction methods are not suitable for oil shale samples.

关 键 词：油页岩 微生物总DNA 提取方法 PCR—DGGE SDS高盐提取法 

分 类 号：TP274[自动化与计算机技术—检测技术与自动化装置]