Clegg C D, Ritz K, Griffiths B S
Cellular and Environmental Physiology Department, Scottish Crop Research Institute, Invergowrie, Dundee, UK.
Lett Appl Microbiol. 1997 Jul;25(1):30-3. doi: 10.1046/j.1472-765x.1997.00166.x.
This paper describes a protocol effective at extracting high yields of high-purity microbial community DNA from humified soils. DNA was extracted from soil by lysozyme, SDS and freeze-thaw lysis, precipitated and then subjected to a double caesium chloride density gradient centrifugation stage before concentrating and washing. Evaluation using three soils yielded up to 30 micrograms DNA g-1 dry soil, with absorbance ratios at 260:230 nm and 260:280 nm of 1.6-2.0. The DNA extracted from the three soils was digested by four restriction enzymes and a 16S rDNA eubacterial product was amplified by PCR. These tests indicated that the DNA obtained by the protocol was sufficiently pure for molecular biological analysis.
本文描述了一种从腐殖化土壤中高效提取高纯度微生物群落DNA的方法。通过溶菌酶、SDS和冻融裂解从土壤中提取DNA，沉淀后在浓缩和洗涤之前进行双氯化铯密度梯度离心步骤。使用三种土壤进行评估，得到了高达30微克DNA/克干土的产量，在260:230纳米和260:280纳米处的吸光度比值为1.6 - 2.0。从这三种土壤中提取的DNA用四种限制性内切酶进行消化，并通过PCR扩增出16S rDNA真细菌产物。这些测试表明，通过该方法获得的DNA纯度足以用于分子生物学分析。
