IMIS | Flanders Marine Institute
 

Flanders Marine Institute

Platform for marine research

IMIS

Publications | Institutes | Persons | Datasets | Projects | Maps
[ report an error in this record ]basket (0): add | show Printer-friendly version

Stable Isotope Probing Combined with Magnetic Bead Capture Hybridization of 16S rRNA: Linking Microbial Community Function to Phylogeny of Sulfate-Reducing Deltaproteobacteria in Marine Sediments
Miyatake, T.; MacGregor, B.J.; Boschker, E. (2009). Stable Isotope Probing Combined with Magnetic Bead Capture Hybridization of 16S rRNA: Linking Microbial Community Function to Phylogeny of Sulfate-Reducing Deltaproteobacteria in Marine Sediments. Appl. Environ. Microbiol. 75: 4927-4935
In: Applied and Environmental Microbiology. American Society for Microbiology: Washington. ISSN 0099-2240, more
Peer reviewed article  

Available in Authors | Dataset 

Keyword
    Marine

Authors  Top | Dataset 
  • Miyatake, T., more
  • MacGregor, B.J.
  • Boschker, E., more

Abstract
    We further developed the stable isotope probing, magnetic bead-capture method to make it applicable for linking microbial community function to phylogeny at the class and family level. The main improvements were a substantial decrease in the protocol blank and an approximately 10 fold increase in detection limit by using a micro-elemental analyzer coupled to isotope ratio mass spectrometry to determine 13C-labeling of isolated 16S rRNA. We demonstrated the method by studying substrate utilization by Desulfobacteraceae, a dominant group of complete oxidizing sulfate-reducing Deltaproteobacteria in marine sediments. Stable-isotope-labeled [13C]glucose, [13C]propionate, or [13C]acetate were fed into an anoxic intertidal sediment. We applied a nested set of three biotin-labeled oligonucleotide probes to capture Bacteria, Deltaproteobacteria and finally Desulfobacteraceae rRNA by using hydrophobic streptavidin-coated paramagnetic beads. Target specificity of the probes was examined with pure cultures of target and non-target species and by determining the phylogenetic composition of the captured sediment rRNA. Specificity of the final protocol was generally very good as more than 90% of the captured 16S rRNA belonged to the target range of the probes. Our results indicated that Desulfobacteraceae were important consumers of propionate but not of glucose. However, results for acetate utilization were less conclusive due to lower and more variable labeling levels in captured rRNA. The main advantage of the method in this study over other nucleic acid based stable isotope probing methods is that 13C-labeling can be much lower, to the extent that even natural abundance {delta}13C ratios can be studied.

Dataset
  • Mag-SIP: Linking Microbial Community Function to Phylogeny of Sulfate-Reducing Deltaproteobacteria in Marine Sediments, more

All data in IMIS is subject to the VLIZ privacy policy Top | Authors | Dataset