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Gin‐mediated site‐specific recombination in bacteriophage Mu DNA: overproduction of the protein and inversion in vitro
Author(s) -
Mertens Gabriele,
Hoffmann Andrea,
Blöcker Helmut,
Frank Ronald,
Kahmann Regine
Publication year - 1984
Publication title -
the embo journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 7.484
H-Index - 392
eISSN - 1460-2075
pISSN - 0261-4189
DOI - 10.1002/j.1460-2075.1984.tb02148.x
Subject(s) - biology , overproduction , bacteriophage , recombination , dna , genetics , inversion (geology) , in vitro , microbiology and biotechnology , escherichia coli , gene , paleontology , structural basin
Inversion of the G segment in bacteriophage Mu DNA occurs by a site‐specific recombination event and determines the host specificity of Mu phage particles produced. Inversion is mediated by a Mu function (Gin). The gin gene has been placed under control of the inducible λ pL promoter and a synthetic Shine‐Dalgarno linker upstream of the initiation codon. The Gin protein content in induced cells is boosted to ˜10% of total protein. Partially purified extracts from overproducing strains promote efficient inversion of the G DNA segment in vitro which is visualized by agarose gel electrophoresis of the substrate DNA after cutting with appropriate restriction endonucleases. The in vitro reaction requires Mg 2+ , a super‐coiled DNA substrate and occurs in the absence of exogenous ATP. Inversion from the G(+) to the G(−) orientation is as efficient as the switch from G(−) to G(+).