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Applied and Environmental Microbiology, May 2008, p. 2902-2907, Vol. 74, No. 9
0099-2240/08/$08.00+0     doi:10.1128/AEM.02161-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Innovative Methods for Soil DNA Purification Tested in Soils with Widely Differing Characteristics{triangledown} ,{dagger}

Marketa Sagova-Mareckova,1* Ladislav Cermak,1 Jitka Novotna,1 Kamila Plhackova,1 Jana Forstova,2 and Jan Kopecky1

Laboratory for Biology of Secondary Metabolism, Institute of Microbiology of the ASCR v.v.i., Videnska 1083, CZ-14220 Prague 4, Czech Republic,1 Institute of Applied Mathematics and Information Technologies, Faculty of Sciences, Charles University in Prague, Albertov 6, CZ-12843 Prague 2, Czech Republic2

Received 21 September 2007/ Accepted 5 March 2008

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 CaCO3 or purification of extracted DNA by CaCl2. 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 CaCl2 purification and in 93% of cases by the method based on CaCO3 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.

* Corresponding author. Mailing address: Crop Research Institute v.v.i., Drnovska 507, CZ-161 06 Prague 6, Czech Republic. Phone: 420 233 022 265. Fax: 420 233 310 636. E-mail: marketa{at}marecek.cz

{triangledown} Published ahead of print on 14 March 2008.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

Applied and Environmental Microbiology, May 2008, p. 2902-2907, Vol. 74, No. 9
0099-2240/08/$08.00+0     doi:10.1128/AEM.02161-07
Copyright © 2008, American Society for Microbiology. All Rights Reserved.



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