Journal of Microbiology and Biotechnology

The Korean Society for Microbiology and Biotechnology (한국미생물·생명공학회)
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Sequence-Based Screening for Putative Polyketide Synthase Gene-Harboring Clones from a Soil Metagenome Library

  • JI SANG CHUN (Korea Research Institute of Bioscience and Biotechnology) ;
  • KIM DOCKYU (Korea Research Institute of Bioscience and Biotechnology) ;
  • YOON JUNG-HOON (Korea Research Institute of Bioscience and Biotechnology) ;
  • OH TAE-KWANG (Korea Research Institute of Bioscience and Biotechnology) ;
  • LEE CHOONG-HWAN (Korea Research Institute of Bioscience and Biotechnology)
  • Published : 2006.01.01
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Abstract

A soil metagenomic library was constructed using an E. coli-fosmid cloning system with environmental DNAs extracted from Kwangreung forest topsoil. We targeted the genes involved in the biosynthesis of bacterial polyketides. Initially, a total of 36 clone pools (10,800 clones) were explored by the PCR-based method using the metagenomic DNAs from each pool and a degenerate primer set, which has been designed based on the highly conserved regions among ketoacyl synthase (KS) domains in actinomycete type I polyketide synthases (PKS Is). Six clone pools were tentatively selected as positive and further examined through a hybridization-based method for selecting a fosmid clone containing PKS I genes. Colony hybridization was performed against fosmid clones from the 6 positive pools, and finally 4 clones were picked out and confirmed to contain the conserved DNA fragment of KS domains. In this study, we present a simple and feasible sorting method for a desired clone from metagenomic libraries.

Keywords

References

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