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Sul, Woo Jun,Park, Joonhong,Quensen III, John F.,Rodrigues, Jorge L. M.,Seliger, Laurie,Tsoi, Tamara V.,Zylstra, Gerben J.,Tiedje, James M. American Society for Microbiology 2009 Applied and environmental microbiology Vol.75 No.17
<B>ABSTRACT</B><P>Stable isotope probing with [<SUP>13</SUP>C]biphenyl was used to explore the genetic properties of indigenous bacteria able to grow on biphenyl in PCB-contaminated River Raisin sediment. A bacterial 16S rRNA gene clone library generated from [<SUP>13</SUP>C]DNA after a 14-day incubation with [<SUP>13</SUP>C]biphenyl revealed the dominant organisms to be members of the genera <I>Achromobacter</I> and <I>Pseudomonas</I>. A library built from PCR amplification of genes for aromatic-ring-hydroxylating dioxygenases from the [<SUP>13</SUP>C]DNA fraction revealed two sequence groups similar to <I>bphA</I> (encoding biphenyl dioxygenase) of <I>Comamonas testosteroni</I> strain B-356 and of <I>Rhodococcus</I> sp. RHA1. A library of 1,568 cosmid clones was produced from the [<SUP>13</SUP>C]DNA fraction. A 31.8-kb cosmid clone, detected by aromatic dioxygenase primers, contained genes of biphenyl dioxygenase subunits <I>bphAE</I>, while the rest of the clone's sequence was similar to that of an unknown member of the <I>Gammaproteobacteria</I>. A discrepancy in G+C content near the <I>bphAE</I> genes implies their recent acquisition, possibly by horizontal transfer. The biphenyl dioxygenase from the cosmid clone oxidized biphenyl and unsubstituted and <I>para</I>-only-substituted rings of polychlorinated biphenyl (PCB) congeners. A DNA-stable isotope probing-based cosmid library enabled the retrieval of functional genes from an uncultivated organism capable of PCB metabolism and suggest dispersed dioxygenase gene organization in nature.</P>