SUMMARY: A 6.5 kb DNA fragment containing a chloramphenicol-resistance gene of ISP5230 was cloned in M252 using the high-copy-number plasmid vector pIJ702. The gene was located within a 2.4 kb I-I fragment of the cloned DNA and encoded an enzyme (chloramphenicol hydrolase) that catalysed removal of the dichloroacetyl moiety from the antibiotic. The deacylated product, -nitrophenylserinol, was metabolized to -nitrobenzyl alcohol and other compounds by enzymes present in M252. Examination of the genomic DNA from several sources using the cloned 6.5 kb I fragment from ISP5230 as a probe showed a hybridizing region in the DNA from 13s but none in the DNA from another chloramphenicol producer, NRRLB 3559. The resistance phenotype was not expressed when the 6.5 kb I fragment or a subfragment was subcloned behind the -promoter of plasmid pTZ18R in


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