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Expression vector with two-step control by the cI-pR-Q-p'R-qut-t'R module of coliphage lambda.
L A Petrenko1, I P Gileva, V V Kravchenko
1All-Union Molecular Biology Research Institute, Koltsovo, Novosibirsk Region, U.S.S.R.
Gene
|May 15, 1989
Summary
This study presents a novel plasmid expression vector, pCEQ3, for high-level protein production in Escherichia coli. It utilizes a temperature-inducible system from bacteriophage lambda for controlled gene expression.
Area of Science:
- Molecular Biology
- Genetic Engineering
- Microbiology
Background:
- Efficient gene expression systems are crucial for biotechnology and research.
- Bacteriophage lambda regulatory elements offer unique control mechanisms for gene transcription.
Purpose of the Study:
- To construct and characterize a novel plasmid expression vector, pCEQ3.
- To enable high-level, temperature-regulated expression of foreign genes in Escherichia coli.
Main Methods:
- Construction of the pCEQ3 vector by replacing a DNA fragment in pBR327 with a lambda phage DNA module.
- Incorporation of the cI857 repressor gene, pR promoter, Q gene, and p'R promoter.
- Insertion of foreign genes at the EcoRI site downstream of p'R.
Main Results:
- The pCEQ3 vector demonstrates temperature-dependent transcriptional regulation.
- Low temperatures (30°C) repress transcription, while a shift to 37°C activates transcription from p'R.
- Expression of cloned genes downstream of p'R results in extremely high product levels in E. coli.
Conclusions:
- The pCEQ3 vector provides a robust and inducible system for high-yield protein expression.
- This system is valuable for applications requiring controlled and amplified production of recombinant proteins.