Innovative methods for soil DNA purification tested in soils with widely differing characteristics.
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.
Sagova-Mareckova
Marketa
M
Crop Research Institute v.v.i., Drnovska 507, CZ-161 06 Prague 6, Czech Republic. marketa@marecek.cz
Cermak
Ladislav
L
Novotna
Jitka
J
Plhackova
Kamila
K
Forstova
Jana
J
Kopecky
Jan
J
D044822:Biodiversity; D002119:Calcium Carbonate; D002122:Calcium Chloride; D004269:DNA, Bacterial / Q000235:genetics / Q000302:isolation & purification*; D006812:Humic Substances; D008967:Molecular Biology / Q000379:methods*; D016133:Polymerase Chain Reaction; D012988:Soil Microbiology
D004269:DNA, Bacterial; D006812:Humic Substances; D002119:Calcium Carbonate; D002122:Calcium Chloride