[A highly effective extraction method for PCR analysis of soil microbial DNA].
In this paper, soil microbial DNA was extracted by cetyltrimethyl ammonium bromide (CTAB) -lysosome-protease K-freezing thaw lysing, and precipitated and purified with PEG 8000. The results showed that this method was simple and effective for the PCR analysis of soil microbial DNA. To effectively remove humid acid, test soil was pre-washed by polyvinylpyrrolidone (PVP) buffer and added with CaCl2 and bovine serum albumin (BSA). The precipitation with PEG8000 could obtain high quality DNA, and the lysing with (CTAB) -lysosome-protease K-freezing thaw could get large fragment DNA. Using this method, the yield of soil microbial DNA under Heptacodium miconioides forest was 9.22 microg x g(-1), with A260/A080 being 1.65. The extracted DNA was proved to be successfully used for further PCR amplification and amplified ribosomal DNA restriction analysis (ARDRA). The optimal template DNA concentration in PCR amplification was 0.67 ng x microl(-1). The establishment of this simple, rapid and effective method made it possible to study the molecular ecology of soil microbes in a large scale.
Li
Junmin
J
Institute of Ecology, Taizhou University, Linhai 317000, China. lijm@mail.tzptt.zj.cn
Jin
Zexin
Z
D004269:DNA, Bacterial / Q000302:isolation & purification*; D004275:DNA, Ribosomal / Q000032:analysis / Q000302:isolation & purification; D005821:Genetic Techniques; D016133:Polymerase Chain Reaction / Q000379:methods*; D012988:Soil Microbiology
D004269:DNA, Bacterial; D004275:DNA, Ribosomal