Use of Pulsed Field Gel Electrophoresis and Transposon Mutagenesis to Estimate the Minimal Number of Genes Required for Motility in Caulobacter crescentus

INVESTIGATIONS

Use of Pulsed Field Gel Electrophoresis and Transposon Mutagenesis to Estimate the Minimal Number of Genes Required for Motility in Caulobacter crescentus

B. Ely and T. W. Ely
Department of Biological Sciences, University of South Carolina, Columbia, South Carolina 29208

To facilitate the mapping of transposon insertion mutations in Caulobacter crescentus, we have used pulsed field gel electrophoresis to construct a detailed physical and genetic map of the C. crescentus genome. Restriction fragments were generated by DraI, AseI, or SpeI which cleave the C. crescentus 40, 13, and 26 times, respectively, and Tn5 insertions were used to align the restriction fragments generated by each of the enzymes. The utility of the resulting map was demonstrated by determining the chromosomal locations of a collection of flagellar mutations. As a result of this study, we were able to identify ten new flagellar genes at various locations on the chromosome. Thus, at least 48 genes are required for the assembly of a functional flagellum in C. crescentus.

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