Overproduction of the toxic protein, bovine pancreatic DNaseI, in Escherichia coli using a tightly controlled T7-promoter-based vector.
A synthetic gene coding for bovine pancreatic DNaseI has been cloned under the control of a T7 promoter present on the plasmid pET11. This construct yields a stable Escherichia coli transformant only when transcription from this promoter is tightly controlled. Production of recombinant DNaseI (reDNaseI) is achieved by infection of the cells with a mutant lambda phage, CE6, which carries the gene encoding T7 RNA polymerase. Induced bacterial cultures yield in excess of 2 mg per litre of reDNaseI after purification.
Doherty
A J
AJ
Department of Biochemistry, University of Southampton, UK.
Connolly
B A
BA
Worrall
A F
AF
D000818:Animals; D017123:Bacteriophage T7 / Q000235:genetics; D002417:Cattle; D003001:Cloning, Molecular; D012321:DNA-Directed RNA Polymerases / Q000235:genetics; D003850:Deoxyribonuclease I / Q000235:genetics*; D004926:Escherichia coli / Q000235:genetics*; D005813:Genes, Synthetic; D010179:Pancreas / Q000201:enzymology; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014764:Viral Proteins
D014764:Viral Proteins; C062210:bacteriophage T7 RNA polymerase; D012321:DNA-Directed RNA Polymerases; D003850:Deoxyribonuclease I