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An improved Eschehchia coli‐Rhodococcus shuttle vector and plasmid transformation in Rhodococcus spp. using electroporation
Author(s) -
Shao Z.,
Dick W.A.,
Behki R.M.
Publication year - 1995
Publication title -
letters in applied microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.698
H-Index - 110
eISSN - 1472-765X
pISSN - 0266-8254
DOI - 10.1111/j.1472-765x.1995.tb01056.x
Subject(s) - rhodococcus , shuttle vector , electroporation , plasmid , transformation (genetics) , thiostrepton , biology , microbiology and biotechnology , rhodococcus equi , dna , escherichia coli , bacteria , strain (injury) , genetics , vector (molecular biology) , gene , recombinant dna , virulence , ribosome , rna , anatomy
The genetic studies of metabolically diverse Rhodococcus spp. have been hampered by the lack of a system of introducing exogenous DNA. The authors improved an existing Escherichia coli‐Rhodococcus shuttle vector (pMVS301) by removing much of the DNA not needed for replication and adding a multicloning site. This improved vector (pBS305) is 7·9 kb in length. Its ability to transform Rhodococcus was tested using electroporation parameters optimized for introduction of pMVS301 into Rhodococcus. Transformation efficiencies as high as 10 5 cfu μg ‐1 DNA were obtained although efficiencies varied depending on the Rhodococcus strain tested. The improved vector pBS305 offers great utility for genetic studies of Rhodococcus because its small size enables movement of large inserts of DNA into Rhodococcus , it has multicloning sites, contains a highly selective thiostrepton marker, and can be replicated in both E. coli and Rhodococcus.