露天矿排土场土壤微生物总DNA提取方法的研究

对露天矿排土场土壤微生物总DNA的提取方法进行了研究,将化学法(SDS高盐法和PBS缓冲液法)、酶法(溶菌酶和蛋白酶K)和机械法(循环冻融)组合为12种细胞裂解方案。琼脂糖凝胶电泳和UV检测的结果表明,SDS高盐法与酶法和机械法组合的6种方案所获得的DNA均具有较好的完整性和较大的浓度。为缩短试验时间,减小DNA在循环冻融过程中造成机械断裂的可能性,故SDS高盐裂解液不经循环冻融、并加入蛋白酶K和溶菌酶的方案组合为最佳,该结果为从分子水平上研究露天矿排土场土壤微生物的多样性及以PCR为基础的研究提供了可行的方法。

Study of Extraction Methods for Soil Microbial Total DNA in the Opencast Coal Mine Dump

Extraction methods for soil microbial total DNA were studied in opencast coal mine dump.The chemical methods（SDS hyperhaline buffer and PBS buffer）,the enzyme methods（lysozyme and proteinase K）and the mechanical method（freeze-thaw cycles）were grouped into twelve kinds of schemes.Agarose gel electrophoresis and UV tests showed that six kinds of methods could extract DNA having better integrity and greater consistency. These methods were made up of SDS hyperhaline buffer,the enzyme methods and the mechanical method. In order to shorten test time and reduce feasibility of DNA fracture,the scheme that was SDS hyperhaline buffer joining proteinase K and lysozyme without mechanical method was the best.The results provided a feasible method for studying soil microbial diversity of opencast coal mine dump in the molecular level.