ArticleThe Journal of biological chemistry2010
Prospecting metagenomic enzyme subfamily genes for DNA family shuffling by a novel PCR-based approach.
Article in The Journal of biological chemistry, 2010. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 11 papers.
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Who cites it
11 citing papers in PubMed.
- Diversity of tryptophan halogenases in sponges of the genus Aplysina.FEMS microbiology ecology · 2019Article
- Concepts and Methods to Access Novel Antibiotics from Actinomycetes.Antibiotics (Basel, Switzerland) · 2018Review
- Molecular Diagnostic for Prospecting Polyhydroxyalkanoate-Producing Bacteria.Bioengineering (Basel, Switzerland) · 2017Article
- Gene-specific amplicons from metagenomes as an alternative to directed evolution for enzyme screening: a case study using phenylacetaldehyde reductases.FEBS open bio · 2016Article
- Culture-independent discovery of natural products from soil metagenomes.Journal of industrial microbiology & biotechnology · 2016Review
- Efficient PCR-based amplification of diverse alcohol dehydrogenase genes from metagenomes for improving biocatalysis: screening of gene-specific amplicons from metagenomes.Applied and environmental microbiology · 2014Article
- Isolation an Aldehyde Dehydrogenase Gene from Metagenomics Based on Semi-nest Touch-Down PCR.Indian journal of microbiology · 2014Article
- New extremophilic lipases and esterases from metagenomics.Current protein & peptide science · 2014Review
- Strategies for the multiplex mapping of genes to traits.Microbial cell factories · 2013Review
- Directed evolution: an evolving and enabling synthetic biology tool.Current opinion in chemical biology · 2012Review
- Biocatalysts and small molecule products from metagenomic studies.Current opinion in chemical biology · 2012Review
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Authors and funding
9 authors.
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Abstract
DNA family shuffling is a powerful method for enzyme engineering, which utilizes recombination of naturally occurring functional diversity to accelerate laboratory-directed evolution. However, the use of this technique has been hindered by the scarcity of family genes with the required level of sequence identity in the genome database. We describe here a strategy for collecting metagenomic homologous genes for DNA shuffling from environmental samples by truncated metagenomic gene-specific PCR (TMGS-PCR). Using identified metagenomic gene-specific primers, twenty-three 921-bp truncated lipase gene fragments, which shared 64-99% identity with each other and formed a distinct subfamily of lipases, were retrieved from 60 metagenomic samples. These lipase genes were shuffled, and selected active clones were characterized. The chimeric clones show extensive functional and genetic diversity, as demonstrated by functional characterization and sequence analysis. Our results indicate that homologous sequences of genes captured by TMGS-PCR can be used as suitable genetic material for DNA family shuffling with broad applications in enzyme engineering.
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