FIG 3.

Transformation frequencies of M. florum oriC plasmids and recombination with the chromosome. (A) Transformation frequencies of M. florum oriC plasmids using polyethylene glycol (PEG)-mediated transformation procedure. o1, pMflT-o1; o2, pMflT-o2; o3, pMflT-o3; o4, pMflT-o4. Error bars indicate the standard deviations calculated from the results of six independent biological replicates. Asterisks indicate transformation frequencies below the detection limit. (B and C) Southern blot analysis of pMflT-o3 (B) and pMflT-o4 (C) recombination with the M. florum chromosome. Fragment sizes corresponding to the integrated and extrachromosomal forms of each plasmid are indicated. I, plasmid integrated at the oriC region of the chromosome; I_A, plasmid integrated at the rpmH-dnaA intergenic region; I_B, plasmid integrated at the dnaA-dnaN intergenic region; E, plasmid as an extrachromosomal element. Twelve isolated M. florum clones were analyzed for each plasmid (clone number indicated above each well). P, purified plasmid control; N, M. florum L1 wild-type (WT) genomic DNA (negative control).