期刊
FRONTIERS IN MICROBIOLOGY
卷 5, 期 -, 页码 -出版社
FRONTIERS MEDIA SA
DOI: 10.3389/fmicb.2014.00189
关键词
metagenomics; functional metagenomics; gut microbiome; microbiota; salt tolerance; BIOLOG; phenotype microarray; transcriptional regulator
类别
资金
- Science Foundation Ireland (SFI) [07/CE/B1368]
- Alimentary Pharmabiotic Centre
- Royal Society
- EU [FP7-PEOPLE-2012-IAPP]
In the current study, a number of salt-tolerant clones previously isolated from a human gut metagenomic library were screened using Phenotype MicroArray (PM) technology to assess their functional capacity. PM's can be used to study gene function, pathogenicity, metabolic capacity and identify drug targets using a series of specialized microtitre plate assays, where each well of the microtitre plate contains a different set of conditions and tests a different phenotype. Cellular respiration is monitored colorimetrically by the reduction of a tetrazolium dye. One clone, SMG 9, was found to be positive for utilization/transport of Lcarnitine (a well-characterized osmoprotectant) in the presence of 6% w/v sodium chloride (NaCI). Subsequent experiments revealed a significant growth advantage in minimal media containing NaCI and Lcarnitine. Fosmid sequencing revealed putative candidate genes responsible for the phenotype. Subsequent cloning of two genes did not replicate the Lcarnitine-associated phenotype, although one of the genes, a a54-dependent transcriptional regulator, did confer salt tolerance to Escherichia coli when expressed in isolation. The original clone, SMG 9, was subsequently found to have lost the original observed phenotype upon further investigation. Nevertheless, this study demonstrates the usefulness of a phenomic approach to assign a functional role to metagenome-derived clones.
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