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DNA-, rRNA- and mRNA-based stable isotope probing of aerobic methanotrophs in lake sediment

DNA-, rRNA- and mRNA-based stable isotope probing of aerobic methanotrophs in lake sediment
DNA-, rRNA- and mRNA-based stable isotope probing of aerobic methanotrophs in lake sediment
A stable isotope probing (SIP) approach was used to study aerobic methane-oxidizing bacteria (methanotrophs) in lake sediment. Oligotrophic Lake Stechlin was chosen because it has a permanently oxic sediment surface. 16S rRNA and the pmoA gene, which encodes a subunit of the methane monooxygenase enzyme, were analysed following the incubation of sediment with 13CH4 and the separation of 13C-labelled DNA and RNA from unlabelled nucleic acids. The incubation with 13CH4 was performed over a 4-day time-course and the pmoA genes and transcripts became progressively labelled such that approximately 70% of the pmoA genes and 80% of the transcripts were labelled at 96 h. The labelling of pmoA mRNA was quicker than pmoA genes, demonstrating that mRNA-SIP is more sensitive than DNA-SIP; however, the general rate of pmoA transcript labelling was comparable to that of the pmoA genes, indicating that the incorporation of 13C into ribonucleic acids of methanotrophs was a gradual process. Labelling of Betaproteobacteria was clearly seen in analyses of 16S rRNA by DNA-SIP and not by RNA-SIP, suggesting that cross-feeding of the 13C was primarily detected by DNA-SIP. In general, we show that the combination of SIP approaches provided valuable information about the activity and growth of the methanotrophic populations and the cross-feeding of methanotroph metabolites by other microorganisms
1462-2920
1153-1167
Dumont, Marc
afd9f08f-bdbb-4cee-b792-1a7f000ee511
Pommerenke, Bianca
b88ad6f7-3df5-46b2-ad72-395ecef447f3
Casper, Peter
90dac29d-fee3-4d8c-8b50-db661e71c80a
Conrad, Ralf
b63adcc7-abe3-4e99-9ce6-20f1cc671d96
Dumont, Marc
afd9f08f-bdbb-4cee-b792-1a7f000ee511
Pommerenke, Bianca
b88ad6f7-3df5-46b2-ad72-395ecef447f3
Casper, Peter
90dac29d-fee3-4d8c-8b50-db661e71c80a
Conrad, Ralf
b63adcc7-abe3-4e99-9ce6-20f1cc671d96

Dumont, Marc, Pommerenke, Bianca, Casper, Peter and Conrad, Ralf (2011) DNA-, rRNA- and mRNA-based stable isotope probing of aerobic methanotrophs in lake sediment. Environmental Microbiology, 13 (5), 1153-1167. (doi:10.1111/j.1462-2920.2010.02415.x).

Record type: Article

Abstract

A stable isotope probing (SIP) approach was used to study aerobic methane-oxidizing bacteria (methanotrophs) in lake sediment. Oligotrophic Lake Stechlin was chosen because it has a permanently oxic sediment surface. 16S rRNA and the pmoA gene, which encodes a subunit of the methane monooxygenase enzyme, were analysed following the incubation of sediment with 13CH4 and the separation of 13C-labelled DNA and RNA from unlabelled nucleic acids. The incubation with 13CH4 was performed over a 4-day time-course and the pmoA genes and transcripts became progressively labelled such that approximately 70% of the pmoA genes and 80% of the transcripts were labelled at 96 h. The labelling of pmoA mRNA was quicker than pmoA genes, demonstrating that mRNA-SIP is more sensitive than DNA-SIP; however, the general rate of pmoA transcript labelling was comparable to that of the pmoA genes, indicating that the incorporation of 13C into ribonucleic acids of methanotrophs was a gradual process. Labelling of Betaproteobacteria was clearly seen in analyses of 16S rRNA by DNA-SIP and not by RNA-SIP, suggesting that cross-feeding of the 13C was primarily detected by DNA-SIP. In general, we show that the combination of SIP approaches provided valuable information about the activity and growth of the methanotrophic populations and the cross-feeding of methanotroph metabolites by other microorganisms

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Published date: May 2011
Organisations: Centre for Biological Sciences, Environmental

Identifiers

Local EPrints ID: 387926
URI: http://eprints.soton.ac.uk/id/eprint/387926
ISSN: 1462-2920
PURE UUID: ad7309b7-2b7d-4034-a9e9-c8ef1c373235
ORCID for Marc Dumont: ORCID iD orcid.org/0000-0002-7347-8668

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Date deposited: 13 Jun 2016 10:50
Last modified: 10 Dec 2019 01:32

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