Temperature-mediated regulation and downstream inducible selection for controlling gene expression from the bacteriophage lambda pL promoter.
We have examined in detail the effects of various induction temperatures on the expression of a heterologous fusion gene controlled by the bacteriophage lambda PL promoter in a heat-inducible Escherichia coli expression system which utilizes the CIts857 repressor. Experiments performed over a temperature range spanning 29-42 degrees C indicate that, under our conditions, temperatures as low as 29 degrees C may be required to fully repress the CI857-controlled transcription from PL, and that the highest protein yields are obtained after induction at 36 degrees C for 6 h. We cloned the cat reporter gene downstream from a heterologous gene controlled by PL and found that cat expression at a low induction temperature permits the monitoring of productive transcription through the heterologous gene and thus aids in selecting transformants that are capable of producing the heterologous protein in E. coli.
Lowman
H B
HB
Department of Chemistry, Purdue University, West Lafayette, IN 47907.
Bina
M
M
D010582:Bacteriophage lambda / Q000235:genetics*; D002702:Chloramphenicol Resistance / Q000235:genetics; D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics; D015967:Gene Expression Regulation, Viral; D005798:Genes, Bacterial; D005814:Genes, Viral; D010957:Plasmids; D011401:Promoter Regions, Genetic; D013696:Temperature