Tightly regulated vectors for the cloning and expression of toxic genes

Journal of Microbiological Methods
Larry AnthonyM S Filutowicz

Abstract

A series of low-copy expression vectors that permits the stable maintenance and regulated expression of highly toxic gene products has been developed. These vectors utilize the lactose promoter/operator system, and protect against read-through transcription from other promoters on the plasmid by placement of the rrnB T1T2 terminators upstream of the lactose promoter. For additional regulatory control, the vectors utilize low-copy origins of replication. Either the pMPP6 origin (pSC101-derived) is used for cloning into Escherichia coli or related species, or the broad-host-range RK2 origin of replication is utilized for cloning into the majority of Gram-negative bacteria. The resulting plasmids have no detectable leaky expression. To test these vectors, the genes for the bacteriocidal colicins D, E3, and E7 were cloned and stably maintained in the absence of their immunity genes. Upon induction with isopropyl-beta-D-thiogalactopyranoside (IPTG), cell death was observed, indicating expression of each colicin. These low-copy expression vectors will be useful for the cloning and expression of toxic genes in bacterial systems.

References

Apr 1, 1979·Proceedings of the National Academy of Sciences of the United States of America·D H Figurski, D R Helinski
Jan 1, 1990·Folia Microbiologica·J SmardaZ Vrbická
May 5, 1986·Journal of Molecular Biology·F W Studier, B A Moffatt
Feb 1, 1984·Journal of General Microbiology·P R Lehrbach, P Broda
Jan 1, 1983·Proceedings of the National Academy of Sciences of the United States of America·H A de BoerM Vasser
Mar 1, 1994·Molecular Microbiology·D ManenL Caro
Oct 1, 1993·Protein Expression and Purification·L H NguyenR R Burgess
Mar 22, 1996·Journal of Molecular Biology·J MüllerB Müller-Hill
Sep 5, 1997·Science·F R BlattnerY Shao
Nov 9, 2002·Biochimie·Haruhiko Masaki, Tetsuhiro Ogawa
Nov 9, 2002·Biochimie·Raz ZarivachMenachem Shoham

❮ Previous
Next ❯

Citations

Mar 7, 2006·Applied and Environmental Microbiology·Eva Vincze, Steve Bowra
Jun 24, 2014·Journal of Computer-aided Molecular Design·Eszter NémethChris Oostenbrink
Jan 22, 2013·Journal of Biological Inorganic Chemistry : JBIC : a Publication of the Society of Biological Inorganic Chemistry·Anikó CzeneBéla Gyurcsik
May 17, 2008·Plasmid·Marcin FilutowiczRavi Shankar
Mar 14, 2013·Letters in Applied Microbiology·N Nakashima, T Tamura
Jun 2, 2015·Biotechnology Advances·Amit Kumar ChaudharyEun Yeol Lee
Feb 6, 2014·Veterinary Microbiology·Chetan V JawaleJohn Hwa Lee
May 6, 2016·Biotechnology Letters·Yasuhiro FujinoKatsumi Doi
Sep 3, 2014·Metallomics : Integrated Biometal Science·Anikó CzeneBéla Gyurcsik
Mar 1, 2007·Biochemistry and Molecular Biology Education : a Bimonthly Publication of the International Union of Biochemistry and Molecular Biology·Brent L NielsenChin-Yo Lin
Oct 28, 2005·Nature·Conrad J HoskinCraig Moritz
May 23, 2013·Acta Crystallographica. Section F, Structural Biology and Crystallization Communications·Anikó CzeneKyosuke Nagata
Dec 31, 2019·Molecular Systems Biology·Belén CallesVíctor de Lorenzo

❮ Previous
Next ❯

Related Concepts

Related Feeds

Aminoglycosides (ASM)

Aminoglycoside is a medicinal and bacteriologic category of traditional Gram-negative antibacterial medications that inhibit protein synthesis and contain as a portion of the molecule an amino-modified glycoside. Discover the latest research on aminoglycoside here.

Aminoglycosides

Aminoglycoside is a medicinal and bacteriologic category of traditional Gram-negative antibacterial medications that inhibit protein synthesis and contain as a portion of the molecule an amino-modified glycoside. Discover the latest research on aminoglycoside here.