The use of the plasmid pHA10 in the isolation of lambda PL promoter mutations

The plasmid pHA10 provides a good positive selection system for PL promoter mutations. It contains the lambda EcoR1-D fragment cloned into the pBR322 plasmid. Inactivation of the temperature sensitive lambda repressor at 42 degrees C leads to transcription of the kil gene and host death. One group of survivors at 42 degrees C is saved due to mutations in the PL promoter. After several screening procedures one such mutant was isolated. Restriction enzyme analysis of the plasmid DNA followed by sequence determination showed it to be a 24 base pair deletion beginning half-way through the "Pribnow box" continuing through part of the -35 region. It removes the OL1 section of the OL operator plus one base pair of OL2. This mutation (called dpl) may be useful to the study of the lambda OLPL regulation system. The effect of base changes within the PL promoter on the promoter activity are discussed.