Learn more: PMC Disclaimer | PMC Copyright Notice
Recovery of DNA from soils and sediments.
Abstract
Experiments were performed to evaluate the effectiveness of two different methodological approaches for recovering DNA from soil and sediment bacterial communities: cell extraction followed by lysis and DNA recovery (cell extraction method) versus direct cell lysis and alkaline extraction to recover DNA (direct lysis method). Efficiency of DNA recovery by each method was determined by spectrophotometric absorbance and using a tritiated thymidine tracer. With both procedures, the use of polyvinylpolypyrrolidone was important for the removal of humic compounds to improve the purity of the recovered DNA; without extensive purification, various restriction enzymes failed to cut added target DNA. Milligram quantities of high-purity DNA were recovered from 100-g samples of both soils and sediments by the direct lysis method, which was a greater than 1-order-of-magnitude-higher yield than by the cell extraction method. The ratio of labeled thymidine to total DNA, however, was higher in the DNA recovered by the cell extraction method. than by the direct lysis method, suggesting that the DNA recovered by the cell extraction method came primarily from active bacterial cells, whereas that recovered by the direct lysis method may have contained DNA from other sources.
Full text
Full text is available as a scanned copy of the original print version. Get a printable copy (PDF file) of the complete article (1.8M), or click on a page image below to browse page by page. Links to PubMed are also available for Selected References.
Images in this article
Selected References
These references are in PubMed. This may not be the complete list of references from this article.
- Atlas RM, Sayler GS. Tracking microorganisms and genes in the environment. Basic Life Sci. 1988;45:31–45. [PubMed] [Google Scholar]
- Bakken LR. Separation and purification of bacteria from soil. Appl Environ Microbiol. 1985 Jun;49(6):1482–1487. [PMC free article] [PubMed] [Google Scholar]
- Balkwill DL, Labeda DP, Casida LE., Jr Simplified procedures for releasing and concentrating microorganisms from soil for transmission electron microscopy viewing as thin-sectioned and frozen-etched preparations. Can J Microbiol. 1975 Mar;21(3):252–262. [PubMed] [Google Scholar]
- Dani GM, Zakian VA. Mitotic and meiotic stability of linear plasmids in yeast. Proc Natl Acad Sci U S A. 1983 Jun;80(11):3406–3410. [PMC free article] [PubMed] [Google Scholar]
- Giovannoni SJ, DeLong EF, Olsen GJ, Pace NR. Phylogenetic group-specific oligodeoxynucleotide probes for identification of single microbial cells. J Bacteriol. 1988 Feb;170(2):720–726. [PMC free article] [PubMed] [Google Scholar]
- Hobbie JE, Daley RJ, Jasper S. Use of nuclepore filters for counting bacteria by fluorescence microscopy. Appl Environ Microbiol. 1977 May;33(5):1225–1228. [PMC free article] [PubMed] [Google Scholar]
- Holben William E, Jansson Janet K, Chelm Barry K, Tiedje James M. DNA Probe Method for the Detection of Specific Microorganisms in the Soil Bacterial Community. Appl Environ Microbiol. 1988 Mar;54(3):703–711. [PMC free article] [PubMed] [Google Scholar]
- McCoy WF, Olson BH. Fluorometric determination of the DNA concentration in municipal drinking water. Appl Environ Microbiol. 1985 Apr;49(4):811–817. [PMC free article] [PubMed] [Google Scholar]
- Paul John H, Deflaun Mary F, Jeffrey Wade H, David Andrew W. Seasonal and Diel Variability in Dissolved DNA and in Microbial Biomass and Activity in a Subtropical Estuary. Appl Environ Microbiol. 1988 Mar;54(3):718–727. [PMC free article] [PubMed] [Google Scholar]
- Paul JH, Jeffrey WH, DeFlaun MF. Dynamics of extracellular DNA in the marine environment. Appl Environ Microbiol. 1987 Jan;53(1):170–179. [PMC free article] [PubMed] [Google Scholar]
- Roszak DB, Colwell RR. Survival strategies of bacteria in the natural environment. Microbiol Rev. 1987 Sep;51(3):365–379. [PMC free article] [PubMed] [Google Scholar]