Figure 9.
Genetic testing of the essentiality of RNase H1 endonuclease activity. A schematic representation of the chromosomal attB locus in strain Mgm4083, which has chromosomal deletions of rnhC and rnhA and a copy of rnhC at the attB locus on an integrated plasmid element conferring kanamycin resistance. Because of spontaneous excision of attB integrated plasmids, replacement with a transfected plasmid carrying a streptomycin resistance gene can be efficiently achieved, allowing swapping of the wild type rnhC gene for wild type and mutated versions of rnhC or rnhA. The results of these marker exchange experiments are given in the table as: (i) the number of strepR colonies recovered after transfection with 50 ng of the plasmid carrying the indicated allele and (ii) the results of genotyping of survivors to confirm allelic exchange.