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

Environ Microbiol. 2011 May;13(5):1153-67. doi: 10.1111/j.1462-2920.2010.02415.x. Epub 2011 Jan 24.


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 (13) CH(4) and the separation of (13) C-labelled DNA and RNA from unlabelled nucleic acids. The incubation with (13) CH(4) 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 (13) C 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 (13) C 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.

MeSH terms

  • Carbon Isotopes / analysis
  • DNA, Bacterial / chemistry
  • DNA, Bacterial / genetics
  • Geologic Sediments / microbiology*
  • Methane / metabolism*
  • Methylococcaceae / classification*
  • Methylococcaceae / genetics
  • Methylococcaceae / metabolism
  • Nucleic Acid Probes / chemistry
  • Oxygenases / genetics
  • Phylogeny
  • RNA, Messenger / chemistry
  • RNA, Messenger / genetics
  • RNA, Ribosomal, 16S / chemistry
  • RNA, Ribosomal, 16S / genetics
  • Sequence Analysis, DNA
  • Time Factors
  • Water Microbiology*


  • Carbon Isotopes
  • DNA, Bacterial
  • Nucleic Acid Probes
  • RNA, Messenger
  • RNA, Ribosomal, 16S
  • Oxygenases
  • methane monooxygenase
  • Methane