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Filamentous Coliphage M13 As A Cloning Vehicle: Insertion Of A HindII Fragment Of The Lac Regulatory Region In M13 Replicative Form In Vitro.

J. Messing, B. Gronenborn, B. Müller-Hill, P. Hans Hopschneider
Published 1977 · Biology, Medicine

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A HindII restriction fragment comprising the Escherichia coli lac regulatory region and the genetic information for the alpha peptide of beta-galactosidase (beta-D-galactosidegalactohydrolase, EC. 3.2.1.23) has been inserted into 1 of the 10 Bsu I cleavage sites of M13 by blunt end ligation. A stable hybrid phage was isolated and identified by its ability to complement the lac alpha function. Further characterization of the hybrid phage includes retransformation studies, agarose gel electrophoresis, DNA-DNA hybridization, and heteroduplex mapping. The insertion point has been localized at 0.083 map unit on thewild-type circular map-i.e., within the intergenic region. The results prove that part of the intergenic region is nonessential and that the phage can be used as a cloning vehicle.
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