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© Research
Publication : FEMS microbiology letters

Low-copy-number T7 vectors for selective gene expression and efficient protein overproduction in Escherichia coli

Scientific Fields
Diseases
Organisms
Applications
Technique

Published in FEMS microbiology letters - 15 Oct 1994

Dersch P, Fsihi H, Bremer E

Link to Pubmed [PMID] – 7988888

FEMS Microbiol. Lett. 1994 Oct;123(1-2):19-26

A set of low-copy-number vectors (pPD) has been constructed that permit selective gene expression and high-level protein overproduction in Escherichia coli, based on the bacteriophage T7 RNA polymerase/T7 promoter system. These plasmids carry a chloramphenicol resistance gene (cat) as a selective marker and an extended multiple cloning site for convenient gene cloning. Their replication is mediated by ori sequences derived from the low-copy-number vector pSC101. The efficient T7 gene 10 promoter present on these vectors allows selective and high-level transcription of cloned genes carrying their own translational initiation signals. In addition, low-copy-number T7 vectors were constructed that permit expression of genes lacking their own transcription and translation initiation elements by providing a ribosome binding site, an ATG start codon and a multiple cloning site devised for the cloning in all three reading frames. The pPD expression vectors were used to achieve high-level overproduction of the E. coli integral outer membrane protein Tsx, and the cytoplasmic enzymes beta-galactosidase (beta Gal) and UTP:alpha-D-glucose-1-phosphate uridylyltransferase (GalU). The characteristics of these low-copy-number T7 expression vectors should prove very useful for the cloning and high-level overexpression of genes whose gene products are deleterious to the E. coli host.

http://www.ncbi.nlm.nih.gov/pubmed/7988888