Isolation of mutations of the phage Mu ner gene.

A method was devised which allows the easy detection of mutations within the ner gene of Mu DNA. This method is based upon the observation that a transcriptional gene A-galK fusion containing the complete ner gene and the cts62 allele does not express the galK gene in an Escherichia coli strain lacking functional integration host factor under inducing conditions (white colonies on MacConkey galactose plates at 42 degrees) In contrast, a gene ner-galK fusion which lacks part of the ner gene exhibits GalK activity (red colonies) on MacConkey galactose plates at 42 degrees. After mutagenesis of a plasmid carrying a transcriptional gene A-galK fusion, putative ner mutants could be identified on indicator plates. Cloning experiments locate the mutation(s) to the right of the HindIII site which is situated within the early promoter of Mu DNA. One of the mutants was sequenced and revealed two substitutions: one within the-10 region of the early promoter, and another near the end of the ner gene. The former lesion was shown to be pleiotropic.