Cloning and complementation analysis of the “Frizzy” genes of Myxococcus xanthus

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Abstract
Fruiting-body formation in Myxococcus xanthus involves the aggregation of cells into raised mounds, where they sporulate. “Frizzy” mutants fail to aggregate into mounds, but rather aggregate into “frizzy” filaments (D.R. Zusman 1982). The frizzy mutations (frz) were found to be genetically linked. The region of DNA carrying the frz genes was cloned in Escherichia coli by selecting for the kanamycin resistance element present on a transposon Tn5 insertion linked to the frz genes. Phage P1 mediated transduction of the cloned DNA into M. xanthus frizzy mutants showed that the cloned DNA could complement the frz mutations. The cloned DNA was analyzed by isolating and characterizing new Tn5 insertions at short intervals within the M. xanthus DNA and by constructing in vitro deletions. The mutated DNA was then transduced into M. xanthus where the cloned DNA became integrated into the bacterial chromosome as gene replacements or as merodiploids. The gene replacement strains allowed us to define the limits of the frz region, since Tn5 insertions in the frz genes resulted in the frizzy phenotype. The merodiploid strains allowed us to perform complementation analyses. Using appropriate crosses, we were able to identify 5–6 frz complementation groups on 7.5 kb of cloned DNA. One of the complementation groups was separated from the others by 1.4 kb of DNA, whereas the others were contiguous. The different frz loci behave as separate transcriptional groups although interactions between some of the gene products are indicated.