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Comparative Study
. 2008 May;74(9):2902-7.
doi: 10.1128/AEM.02161-07. Epub 2008 Mar 14.

Innovative methods for soil DNA purification tested in soils with widely differing characteristics

Marketa Sagova-Mareckova  1 , Ladislav Cermak, Jitka Novotna, Kamila Plhackova, Jana Forstova, Jan Kopecky
Affiliations
Comparative Study

Innovative methods for soil DNA purification tested in soils with widely differing characteristics

Marketa Sagova-Mareckova et al. Appl Environ Microbiol. 2008 May.

Abstract

Seven methods of soil DNA extraction and purification were tested in a set of 14 soils differing in bedrock, texture, pH, salinity, moisture, organic matter content, and vegetation cover. The methods introduced in this study included pretreatment of soil with CaCO(3) or purification of extracted DNA by CaCl(2). The performance of innovated methods was compared to that of the commercial kit Mo Bio PowerSoil and the phenol-chloroform-based method of D. N. Miller, J. E. Bryant, E. L. Madsen, and W. C. Ghiorse (Appl. Environ. Microbiol. 65:4715-4724, 1999). This study demonstrated significant differences between the tested methods in terms of DNA yield, PCR performance, and recovered bacterial diversity. The differences in DNA yields were correlated to vegetation cover, soil pH, and clay content. The differences in PCR performances were correlated to vegetation cover and soil pH. The innovative methods improved PCR performance in our set of soils, in particular for forest acidic soils. PCR was successful in 95% of cases by the method using CaCl(2) purification and in 93% of cases by the method based on CaCO(3) pretreatment, but only in 79% by Mo Bio PowerSoil, for our range of soils. Also, the innovative methods recovered a higher percentage of actinomycete diversity from a subset of three soils. Recommendations include the assessment of soil characteristics prior to selecting the optimal protocol for soil DNA extraction and purification.

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Figures

FIG. 1.
FIG. 1.
(A) DNA yield averages (μg·g−1) (dry weight) of soil from sites with different vegetation cover. (For respective standard deviations of the three measurements, see Table S1 in the supplemental material.) Abbreviations (Methods) stand for DNA extraction methods (see Materials and Methods). (B) Numbers of bacteria in different types of vegetation cover. Data are from direct counts under a microscope.
FIG. 2.
FIG. 2.
PCR performance. Evaluation of PCR success from three independent reactions. Sites are ordered according to pH (low pH at the top and high pH at the bottom). Abbreviations stand for DNA extraction methods (see Materials and Methods).
See this image and copyright information in PMC

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