DNA recovery from soils of diverse composition.

Applied and Environmental Microbiology
J ZhouJ M Tiedje

Abstract

A simple, rapid method for bacterial lysis and direct extraction of DNA from soils with minimal shearing was developed to address the risk of chimera formation from small template DNA during subsequent PCR. The method was based on lysis with a high-salt extraction buffer (1.5 M NaCl) and extended heating (2 to 3 h) of the soil suspension in the presence of sodium dodecyl sulfate (SDS), hexadecyltrimethylammonium bromide, and proteinase K. The extraction method required 6 h and was tested on eight soils differing in organic carbon, clay content, and pH, including ones from which DNA extraction is difficult. The DNA fragment size in crude extracts from all soils was > 23 kb. Preliminary trials indicated that DNA recovery from two soils seeded with gram-negative bacteria was 92 to 99%. When the method was tested on all eight unseeded soils, microscopic examination of indigenous bacteria in soil pellets before and after extraction showed variable cell lysis efficiency (26 to 92%). Crude DNA yields from the eight soils ranged from 2.5 to 26.9 micrograms of DNA g-1, and these were positively correlated with the organic carbon content in the soil (r = 0.73). DNA yields from gram-positive bacteria from pure cultures were two to six tim...Continue Reading

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Citations

Apr 1, 2000·Applied and Environmental Microbiology·F E LöfflerJ M Tiedje
Mar 17, 2000·FEMS Microbiology Ecology·B C Crump, J A Baross
Apr 25, 2001·Applied and Environmental Microbiology·P Y Cheung, B K Kinkle
Apr 25, 2001·Applied and Environmental Microbiology·F Martin-LaurentG Catroux
Sep 26, 2001·Applied and Environmental Microbiology·R A HurtJ Zhou
Oct 9, 2002·Environmental Microbiology·Achim QuaiserChrista Schleper
May 8, 2003·Biological Procedures Online·C YeatesD A Veal
Apr 5, 2003·Applied and Environmental Microbiology·Byron C CrumpJohn E Hobbie
Jul 2, 2003·The Journal of General and Applied Microbiology·Shaila KabirKiminori Itoh
Mar 17, 2004·Journal of Environmental Science and Health. Part. B, Pesticides, Food Contaminants, and Agricultural Wastes·Yong-Hua YangMeng-Cheng Wang
Oct 7, 2004·Applied and Environmental Microbiology·Massimiliano CardinaleDaniele Daffonchio
Aug 13, 2004·Environmental Microbiology·Alexander H TreuschChrista Schleper
Jan 28, 2005·Biotechnology and Bioengineering·Siqing XiaXingmao Ma
Jun 3, 2005·Nature Reviews. Microbiology·Rolf Daniel
Sep 15, 2005·Environmental Microbiology·David A WalshW Ford Doolittle
May 17, 2005·Environmental Microbiology·Hans-Peter GrossartThorsten Brinkhoff
Oct 6, 2005·Applied and Environmental Microbiology·Lynn L WilliamsonJo Handelsman
Dec 8, 2005·Applied and Environmental Microbiology·He Kyoung LimSeon-Woo Lee
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Mar 6, 2008·Electrophoresis·Davorka SutlovicSimun Andjelinovic

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