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Title

Heterologous viral expression systems in fosmid vectors increase the functional analysis potential of metagenomic libraries

AuthorsTerrón-González, Laura ; Medina, Carlos ; Limón-Mortés, M. C.; Santero, Eduardo
Issue Date2013
PublisherNature Publishing Group
CitationScientific Reports 3: 1107 (2013)
AbstractThe extraordinary potential of metagenomic functional analyses to identify activities of interest present in uncultured microorganisms has been limited by reduced gene expression in surrogate hosts. We have developed vectors and specialized E. coli strains as improved metagenomic DNA heterologous expression systems, taking advantage of viral components that prevent transcription termination at metagenomic terminators. One of the systems uses the phage T7 RNA-polymerase to drive metagenomic gene expression, while the other approach uses the lambda phage transcription anti-termination protein N to limit transcription termination. A metagenomic library was constructed and functionally screened to identify genes conferring carbenicillin resistance to E. coli. The use of these enhanced expression systems resulted in a 6-fold increase in the frequency of carbenicillin resistant clones. Subcloning and sequence analysis showed that, besides β-lactamases, efflux pumps are not only able contribute to carbenicillin resistance but may in fact be sufficient by themselves to convey carbenicillin resistance.
DescriptionThis work is licensed under a Creative Commons Attribution-NonCommercial-NoDerivs 3.0 Unported License.
Publisher version (URL)http://dx.doi.org/10.1038/srep01107
URIhttp://hdl.handle.net/10261/129156
DOI10.1038/srep01107
ISSN2045-2322
Appears in Collections:(CABD) Artículos
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