Isolation of a dam ts mutant.

By use of PCR, I will attempt to isolate a temperature sensitive dam mutant. The dam gene will be PCR amplified under conditions that promotes misincorporation, thereby creating random mutations in the dam gene. The wild type dam gene, carried on a low copy plasmid, will then be replaced by the mutated copy and clones will be screened for a dam ts phenotype. This mutant will be very usefull in continuing the study of minichromosome incompatibility in Dam deficient cells.