M. Krsek et Emh. Wellington, Comparison of different methods for the isolation and purification of total community DNA from soil, J MICROB M, 39(1), 1999, pp. 1-16
The efficiency and reproducibility of DNA extraction from soil was tested f
or variations in lyric and purification treatments and their effect on yiel
d and purity of DNA. The extraction yield was improved by increasing the co
ncentration of EDTA or monovalent ions in isolation buffers,, by the introd
uction of mechanical lysis treatments, and by the use of ethanol precipitat
ion in place of PEG precipitation. Purity was improved using buffers with d
ecreasing concentration of EDTA or by reducing the ionic strength of the bu
ffer, and by all mechanical treatments. No lyric treatment was efficient on
its own, the highest purity was achieved using Crombach buffer and a combi
nation of bead-beating with lysozyme and SDS lysis followed by potassium ac
etate and PEG precipitation, phenol/chloroform purification, isopropanol pr
ecipitation, and spermine-HCl precipitation. Sonication sheared the DNA mor
e than bead-heating. Lysozyme and SDS lysis without any mechanical treatmen
ts allowed isolation of larger fragments (40-90 kb). Denaturing gradient ge
l electrophoresis analysis of DNA isolated using a range of lyric treatment
s revealed alterations in band patterns which might reflect differences in
the efficiency of lytic treatments. (C) 1999 Elsevier Science B.V. All righ
ts reserved.