Extraction of mRNA from Soil

被引：88

作者：

Mettel, Carsten ^{[1
]}

Kim, Yongkyu ^{[1
]}

Shrestha, Pravin Malla ^{[1
]}

Liesack, Werner ^{[1
]}

机构：

[1] Max Planck Inst Terr Mikrobiol, D-35043 Marburg, Germany

来源：

APPLIED AND ENVIRONMENTAL MICROBIOLOGY | 2010年 / 76卷 / 17期

关键词：

POLYMERASE CHAIN-REACTION; 16S RIBOSOMAL-RNA; GENE-EXPRESSION; ATMOSPHERIC METHANE; HUMIC SUBSTANCES; RAPID METHOD; DNA; COMMUNITY; SEDIMENTS; PROTOCOL;

D O I：

10.1128/AEM.03047-09

中图分类号：

Q81 [生物工程学（生物技术）]; Q93 [微生物学];

学科分类号：

071005 ; 0836 ; 090102 ; 100705 ;

摘要：

Here, we report an efficient method for extracting high-quality mRNA from soil. Key steps in the isolation of total RNA were low-pH extraction (pH 5.0) and Q-Sepharose chromatography. The removal efficiency of humic acids was 94 to 98% for all soils tested. To enrich mRNA, subtractive hybridization of rRNA was most efficient. Subtractive hybridization may be followed by exonuclease treatment if the focus is on the analysis of unprocessed mRNA. The total extraction method can be completed within 8 h, resulting in enriched mRNA ranging from 200 bp to 4 kb in size.

引用

页码：5995 / 6000

页数：6

共 34 条

[1]

Albert B., 1994, MOL BIOL CELL

[2] Bayesian detection of periodic mRNA time profiles without use of training examples [J].

Andersson, CR ;

Isaksson, A ;

Gustafsson, MG .

BMC BIOINFORMATICS, 2006, 7 (1)

[3] mRNA extraction and reverse transcription-PCR protocol for detection of nifH gene expression by Azotobacter vinelandii in soil [J].

Bürgmann, H ;

Widmer, F ;

Sigler, WV ;

Zeyer, J .

APPLIED AND ENVIRONMENTAL MICROBIOLOGY, 2003, 69 (04) :1928-1935

[4] Initiation of RNA decay in Escherichia coli by 5′ pyrophosphate removal [J].

Celesnik, Helena ;

Deana, Atilio ;

Belasco, Joel G. .

MOLECULAR CELL, 2007, 27 (01) :79-90

[5] Identification of active methanotrophs in a landfill cover soil through detection of expression of 16S rRNA and functional genes [J].

Chen, Yin ;

Dumont, Marc G. ;

Cebron, Aurelie ;

Murrell, J. Colin .

ENVIRONMENTAL MICROBIOLOGY, 2007, 9 (11) :2855-2869

[6] Simple and rapid method for direct extraction of microbial DNA from soil for PCR [J].

Cullen, DW ;

Hirsch, PR .

SOIL BIOLOGY & BIOCHEMISTRY, 1998, 30 (8-9) :983-993

[7] Removal of humic substances from soil DNA using aluminium sulfate [J].

Dong, Dexian ;

Yan, An ;

Liu, Haiming ;

Zhang, Xuehong ;

Xu, Yuquan .

JOURNAL OF MICROBIOLOGICAL METHODS, 2006, 66 (02) :217-222

[8] Comparative analysis of genetic diversity and expression of amoA in wastewater treatment processes [J].

Ebie, Y ;

Noda, N ;

Miura, H ;

Matsumura, M ;

Tsuneda, S ;

Hirata, A ;

Inamori, Y .

APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, 2004, 64 (05) :740-744

[9] Microbial community gene expression in ocean surface waters [J].

Frias-Lopez, Jorge. ;

Shi, Yanmei ;

Tyson, Gene W. ;

Coleman, Maureen L. ;

Schuster, Stephan C. ;

Chisholm, Sallie W. ;

DeLong, Edward F. .

PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 2008, 105 (10) :3805-3810

[10] Detection of Large Numbers of Novel Sequences in the Metatranscriptomes of Complex Marine Microbial Communities [J].

Gilbert, Jack A. ;

Field, Dawn ;

Huang, Ying ;

Edwards, Rob ;

Li, Weizhong ;

Gilna, Paul ;

Joint, Ian .

PLOS ONE, 2008, 3 (08)

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