Comparison of different methods for the isolation and
purification of total community DNA from soil

J 1999

Comparison of different methods for the isolation and purification of total community DNA from soil

KRSEK, Martin and EMH WELLINGTON

Basic information

Original name

Comparison of different methods for the isolation and purification of total community DNA from soil

Authors

KRSEK, Martin and EMH WELLINGTON

Edition

Journal of Microbiological Methods, Amsterdam, Elsevier, 1999, 0167-7012

Other information

Language

English

Type of outcome

Article in a journal

Confidentiality degree

is not subject to a state or trade secret

Impact factor

Impact factor: 1.303

DOI

http://dx.doi.org/10.1016/S0167-7012(99)00093-7

UT WoS

000083660900001

Keywords in English

DNA extraction; microbial community analysis; soil microbiology; total community DNA

Changed: 8/1/2016 11:13, doc. Ing. Martin Krsek, CSc., MSc

Abstract

V originále

The efficiency and reproducibility of DNA extraction from soil was tested for variations in lyric and purification treatments and their effect on yield and purity of DNA. The extraction yield was improved by increasing the concentration of EDTA or monovalent ions in isolation buffers,, by the introduction of mechanical lysis treatments, and by the use of ethanol precipitation in place of PEG precipitation. Purity was improved using buffers with decreasing concentration of EDTA or by reducing the ionic strength of the buffer, and by all mechanical treatments. No lyric treatment was efficient on its own, the highest purity was achieved using Crombach buffer and a combination of bead-beating with lysozyme and SDS lysis followed by potassium acetate and PEG precipitation, phenol/chloroform purification, isopropanol precipitation, and spermine-HCl precipitation. Sonication sheared the DNA more than bead-heating. Lysozyme and SDS lysis without any mechanical treatments allowed isolation of larger fragments (40-90 kb). Denaturing gradient gel electrophoresis analysis of DNA isolated using a range of lyric treatments revealed alterations in band patterns which might reflect differences in the efficiency of lytic treatments. (C) 1999 Elsevier Science B.V. All rights reserved.

Cite

KRSEK, Martin and EMH WELLINGTON. Comparison of different methods for the isolation and purification of total community DNA from soil. Journal of Microbiological Methods. Amsterdam: Elsevier, 1999, vol. 39, No 1, p. 1-16. ISSN 0167-7012. Available from: https://dx.doi.org/10.1016/S0167-7012(99)00093-7.

@article{1323296,
   author = {Krsek, Martin and Wellington, EMH},
   article_location = {Amsterdam},
   article_number = {1},
   doi = {http://dx.doi.org/10.1016/S0167-7012(99)00093-7},
   keywords = {DNA extraction; microbial community analysis; soil microbiology; total community DNA},
   language = {eng},
   issn = {0167-7012},
   journal = {Journal of Microbiological Methods},
   title = {Comparison of different methods for the isolation and purification of total community DNA from soil},
   volume = {39},
   year = {1999}
}

TY  - JOUR
ID  - 1323296
AU  - Krsek, Martin - Wellington, EMH
PY  - 1999
TI  - Comparison of different methods for the isolation and purification of total community DNA from soil
JF  - Journal of Microbiological Methods
VL  - 39
IS  - 1
SP  - 1-16
EP  - 1-16
PB  - Elsevier
SN  - 01677012
KW  - DNA extraction
KW  - microbial community analysis
KW  - soil microbiology
KW  - total community DNA
N2  - The efficiency and reproducibility of DNA extraction from soil was tested for variations in lyric and purification treatments and their effect on yield and purity of DNA. The extraction yield was improved by increasing the concentration of EDTA or monovalent ions in isolation buffers,, by the introduction of mechanical lysis treatments, and by the use of ethanol precipitation in place of PEG precipitation. Purity was improved using buffers with decreasing concentration of EDTA or by reducing the ionic strength of the buffer, and by all mechanical treatments. No lyric treatment was efficient on its own, the highest purity was achieved using Crombach buffer and a combination of bead-beating with lysozyme and SDS lysis followed by potassium acetate and PEG precipitation, phenol/chloroform purification, isopropanol precipitation, and spermine-HCl precipitation. Sonication sheared the DNA more than bead-heating. Lysozyme and SDS lysis without any mechanical treatments allowed isolation of larger fragments (40-90 kb). Denaturing gradient gel electrophoresis analysis of DNA isolated using a range of lyric treatments revealed alterations in band patterns which might reflect differences in the efficiency of lytic treatments. (C) 1999 Elsevier Science B.V. All rights reserved.
ER  -

KRSEK, Martin and EMH WELLINGTON. Comparison of different methods for the isolation and purification of total community DNA from soil. \textit{Journal of Microbiological Methods}. Amsterdam: Elsevier, 1999, vol.~39, No~1, p.~1-16. ISSN~0167-7012. Available from: https://dx.doi.org/10.1016/S0167-7012(99)00093-7.

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