ArticleJournal of visualized experiments : JoVE2011
Bromodeoxyuridine (BrdU) labeling and subsequent fluorescence activated cell sorting for culture-independent identification of dissolved organic carbon-degrading bacterioplankton.
Article in Journal of visualized experiments : JoVE, 2011. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 3 papers.
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3 citing papers in PubMed, 5 citations in OpenAlex.
- From dividing to dormant: embracing the full activity spectrum for environmental microorganisms.Microbiology and molecular biology reviews : MMBR · 2026Review
- Conventional tobacco products harbor unique and heterogenous microbiomes.Environmental research · 2023Article
- A Review of Methods to Determine Viability, Vitality, and Metabolic Rates in Microbiology.Frontiers in microbiology · 2020Review
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5 authors at 2 institutions in 1 country.
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Abstract
Microbes are major agents mediating the degradation of numerous dissolved organic carbon (DOC) substrates in aquatic environments. However, identification of bacterial taxa that transform specific pools of DOC in nature poses a technical challenge. Here we describe an approach that couples bromodeoxyuridine (BrdU) incorporation, fluorescence activated cell sorting (FACS), and 16S rRNA gene-based molecular analysis that allows culture-independent identification of bacterioplankton capable of degrading a specific DOC compound in aquatic environments. Triplicate bacterioplankton microcosms are set up to receive both BrdU and a model DOC compound (DOC amendments), or only BrdU (no-addition control). BrdU substitutes the positions of thymidine in newly synthesized bacterial DNA and BrdU-labeled DNA can be readily immunodetected. Through a 24-hr incubation, bacterioplankton that are able to use the added DOC compound are expected to be selectively activated, and therefore have higher levels of BrdU incorporation (HI cells) than non-responsive cells in the DOC amendments and cells in no-addition controls (low BrdU incorporation cells, LI cells). After fluorescence immunodetection, HI cells are distinguished and physically separated from the LI cells by fluorescence activated cell sorting (FACS). Sorted DOC-responsive cells (HI cells) are extracted for DNA and taxonomically identified through subsequent 16S rRNA gene-based analyses including PCR, clone library construction and sequencing.
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