Development of a Rhodococcus equi-Escherichia coli plasmid shuttle vector.
Isolates of Rhodococcus equi from pneumonic foals possess an 85- or 90-kb virulence-associated plasmid. A prominent, thermoregulated surface antigen, VapA, encoded by these plasmids is thought to be important in virulence. A 135-kb fragment containing the origin of replication of R. equi strain 103 virulence-associated plasmid (pOTS) was identified, sequenced, and its location identified. A simple R. equi-Escherichia coli shuttle plasmid (pRE-1) derived from the E. coli plasmid pACYC177 and the pOTS ori was developed. The plasmid transformed readily and was stable in either host and expressed kanamycin resistance but not beta-lactamase in R. equi. An improved 5.9-kb vector, pRE-7, was developed from pRE-1 and pBluescript. Subcloning of vapA into the multiple cloning site of the beta-galactosidase gene of pRE-7 resulted in weak expression of the gene both in E. coli and R. equi. The shuttle vector may be useful in examining regulation of virulence gene expression in R. equi.
Zheng
H
H
Department of Pathobiology, University of Guelph, Ontario, Canada.
Tkachuk-Saad
O
O
Prescott
J F
JF
D001426:Bacterial Proteins / Q000096:biosynthesis / Q000235:genetics; D001483:Base Sequence; D003001:Cloning, Molecular; D004269:DNA, Bacterial; D004926:Escherichia coli / Q000235:genetics*; D005822:Genetic Vectors; D008074:Lipoproteins / Q000096:biosynthesis / Q000235:genetics; D008969:Molecular Sequence Data; D010957:Plasmids; D018741:Replication Origin; D016929:Rhodococcus equi / Q000235:genetics* / Q000472:pathogenicity; D017422:Sequence Analysis, DNA; D014774:Virulence; D037521:Virulence Factors
D001426:Bacterial Proteins; D004269:DNA, Bacterial; D008074:Lipoproteins; D037521:Virulence Factors