Testing of primers designed for biphenyl 2,3-dioxygenase а-subunit genes detection in bacteria isolated from contaminated soil

The experiments to test PCR-primer set designed for screening bphAl genes (encoding the а-subunit of biphenyl 2,3-dioxygenase) with DNA template of Gram-positive and Gram-negative bacteria-destructors of biphenyl were done. The bphAl gene region, corresponding to cluster Riske of biphenyl 2,3-dioxygenase was amplified. To amplify extended region of the bphAl gene, comprising fragments encoding the cluster Riske and the enzyme active site, forvard primers were used in combination with the proposed by S. Iwai et al. (2010) reverse primer. Some of tested bacteria had bphAl gene similar to this one of Rhodococcus sp. strian HA99 (AB272986) and strain R04 (DQ403247), were significantly different from the bphAl genes, encoding the а-subunit of biphenyl/toluene dioxygenases subfamily. We defined the primers pair, optimal for screening of bphAl genes, encoding of biphenyl 2,3 dioxygenase а-subunits, relating to different subfamilies.