Development of a host-vector system in a Rhodococcus strain and its use for expression of the cloned nitrile hydratase gene cluster

SUMMARY: Two different types of plasmid were isolated from strains of Rhodococcus rhodochrous. Two plasmids, of the same type but from different strains, were combined with Escherichia coli plasmids carrying antibiotic resistance markers to develop E. coli-Rhodocdccus shuttle vectors. The ampicillin and kanamycin resistance markers served for selection in Rhodococcus. Electroporation was used to introduce recombinant plasmid DNA into R. rhodochrous ATCC 12674 at a frequency of 5 × 107transformants per μg DNA. With these host-vector and transformation systems, the nitrile hydratase and amidase genes of a Rhodococcus strain were introduced into the host strain and were efficiently expressed.