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Construction and characterization of a plasmid coding for a fragment of the Escherichia coli recA protein

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Summary

The E. coli recA gene was cloned from the phage λprecA into the vector pBR313. A plasmid, pJL3, was also isolated by cloning a portion of the recA gene into the vector pBR322. pJL3 coded for a fragment of the recA protein 34 Kd (kilodaltons) in size (compared to 40 Kd for the intact protein). This fragment was antigenically related to the recA protein and its synthesis was subject to the same controls as that of the recA protein. The fragment did not express any detectable recA function. When wild-type cells with pJL3 were treated with nalidixic acid, the 34 Kd fragment and the β-lactamase, made from a gene located downstream from the recA segment, were expressed at very high levels. Moreover, in these cells the rate of synthesis of intact recA protein from the chromosome was inhibited about 2-fold, relative to other chromosomal proteins, when compared to wild-type cells with the pBR322 vector. High level expression of the recA protein fragment and/or the β-lactamase appeared to be lethal. The size of the 34 Kd fragment, taken together with the location of chain-termination codons in pJL3, localizes the regulatory region of the recA gene within 100 base pairs.

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  1. Department of Microbiology, College of Medicine, University of Arizona, 85724, Tucson, Arizona, USA

    John W. Little

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Communicated by A.J. Clark

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Little, J.W. Construction and characterization of a plasmid coding for a fragment of the Escherichia coli recA protein. Molec. Gen. Genet. 177, 13–22 (1979). https://doi.org/10.1007/BF00267248

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  • DOI: https://doi.org/10.1007/BF00267248

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