A novel ColE1::Tn3 plasmid vector that allows direct selection of hybrid clones in E. coli

Gene
L S OzakiY Takagi

Abstract

A plasmid, named pKY2289, consists of a whole ColE1 DNA molecule and a complete ampicillin transposon (Tn3). When induced, E. coli K-12 cells which carry pKY2289 promote synthesis of colicin E1, but they are not immune to colicin E1. Inserting a DNA fragment into the EcoRI or the XmaI site of this plasmid abolishes its ability to produce active colicin E1. Thus, cells carrying one of these in vitro recombinant pKY2289 plasmids are able to form normal colonies in the presence of 0.1 microgram/ml of mitomycin C and 50 microgram/ml of ampicillin, while cells carrying the parental pKY2289 form very tiny colonies under the same conditions. This allows a positive selection for an in vitro recombinant pKY2289 molecule carrying a foreign DNA insertion. The properties of cells carrying the original pKY2289 are described and its potential usefulness as a cloning vehicle is demonstrated by cloning all the EcoRI and XmaI fragments of lambda DNA.

References

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Citations

Jul 1, 1988·Molecular & General Genetics : MGG·S Shiota, H Nakayama
Mar 1, 1981·Proceedings of the National Academy of Sciences of the United States of America·H NomiyamaY Takagi
Jul 1, 1982·Proceedings of the National Academy of Sciences of the United States of America·H Ikenaga, K Saigo
May 1, 1982·Proceedings of the National Academy of Sciences of the United States of America·M YamadaA Nakazawa
Oct 15, 1987·European Journal of Biochemistry·S E Westh HansenA Munch-Petersen
Oct 31, 2002·Critical Reviews in Biotechnology·Young-Jun ChoiByong H Lee
Jun 8, 2018·World Journal of Microbiology & Biotechnology·Tiwa RotchanapreedaJamorn Somana

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