Isolation and characterization of a cryptic plasmid from Erwinia citreus ATCC 31623.
A multiple-copy plasmid pPZG500 (3.8 kb) was isolated from a phytopathogenic bacterium Erwinia citreus ATCC 31623. This is the smallest plasmid so far isolated from the genus Erwinia. The plasmid was partially characterized by a set of restriction enzymes and the unique restriction sites were mapped for HindIII, EcoRI, EcoRV and XBaI, while three sites were found for BglII. Nineteen other enzymes did not cut pPZG500. By deletion analyses minimal regions required for replication (ori) and segregational stability (par) were localized on 1.4 kb EcoRV/BglII and 0.7 kb Bgl/II/EcoRI fragments, respectively. The erythromycin resistance marker (Emr) was cloned into pPZG500 and two plasmid derivatives, pPZG502 and pPZG503, were constructed expressing erythromycin resistance as a good selective marker for recombinant selection in Erw. citreus and Escherichia coli. The segregational stability of both constructed plasmids during 90 generations in E. coli JM109 and Erw. citreus C-4 showed that plasmid pPZG503 lacking the presumptive par region was lost from the population at a higher rate. The results of this study demonstrate that plasmid pPZG500 and derivatives are suitable prerequisites for the construction of useful cloning vector(s) in the genus Erwinia.
Bilić
M
M
PLIVA d. d., Research Institute, Department of Biosynthesis and Biotechnology, Zagreb, Croatia.
Delić
V
V
D004261:DNA Replication; D004262:DNA Restriction Enzymes / Q000378:metabolism; D004885:Erwinia / Q000235:genetics*; D005819:Genetic Markers; D010957:Plasmids / Q000302:isolation & purification* / Q000378:metabolism; D015183:Restriction Mapping
D005819:Genetic Markers; D004262:DNA Restriction Enzymes