[Cloning and orientation of cefoperazone resistance gene on plasmid pFC in E. coli HX88108].
Cefoperazone resistance gene (CPZr) has been cloned from plasmid pFC of E. coli HX88108 using the vector pMB9 (TCr, 5.3 kb). The plasmid pFC DNA was partially digested with Sau3A I, and its 1-2 kb fragments were ligated into BamH I site of vector plasmid pMB9. The recombinant DNA was then transformed into E. coli DH5 prepared using calcium chloride. CPZ resistant bacterial colonies were selected on the agar SOB plates containing CPZ (40 micrograms/ml). The resistance to CPZ could be stably reserved in generation after generation. The recombinant plasmids which encoded CPZ resistance were designated pFL11, pFL25, pFL33, pFL82, pFL86 and pFL102. Rapid small-scale preparation of plasmid and DNA restrication enzyme analysis were used for identification of bacterial colonies. Five plasmids DNA physical maps have been established. Comparison of recombinant plasmids maps with pFC map confirmed that the CPZr gene was oriented between nucleotide no. 3200 bp and no. 4800 + 40 bp of plasmid pFC total sequence. Its molecular weight was about 1.6 kb. There were EcoR I, Sma I and Pvu II sites within CPZr gene.
Liang
R
R
Department of Infectious Disease, First Affiliated Hospital, Chengdu.
Fan
X
X
Shu
X
X
Wu
J
J
Liu
R
R
D002438:Cefoperazone / Q000494:pharmacology*; D018550:Cephalosporin Resistance; D003001:Cloning, Molecular; D004926:Escherichia coli / Q000235:genetics*; D005798:Genes, Bacterial; D019450:Genes, MDR; D008970:Molecular Weight; D010957:Plasmids
D002438:Cefoperazone