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. 1983 Aug;80(15):4775–4779. doi: 10.1073/pnas.80.15.4775

Cyclic AMP-dependent constitutive expression of gal operon: use of repressor titration to isolate operator mutations.

M Irani, L Orosz, S Busby, T Taniguchi, S Adhya
PMCID: PMC384127  PMID: 6308647

Abstract

When the gal operator region is present in a multicopy plasmid it binds to all ("titrates") the gal repressor and "induces" the chromosomal gal operon. To make operator mutations (Oa) with reduced affinity toward the repressor, plasmid DNA was irradiated with UV light and mutant derivatives were isolated that were unable to release the chromosomal gal genes from repression. Then with such an Oa plasmid operator revertants were isolated that had reacquired the ability to release repression. Both sets of mutations have been localized by DNA sequence analysis. When the Oa mutations were transferred from the plasmid to the chromosome by recombination these mutant operators were found to make gal expression constitutive (independent of repressor) but still dependent on cAMP, whereas the previously reported gal operator mutants (Oc) are constitutive both in the presence and in the absence of cAMP. The titration method of isolating mutants enables the isolation of strains with operator mutations that also affect normal promoter activity, and it provides an easy way to isolate revertants of operator mutations.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

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