A family of six flagellin genes contributes to the Caulobacter crescentus flagellar filament.

The Caulobacter crescentus flagellar filament is assembled from multiple flagellin proteins that are encoded by six genes. The amino acid sequences of the FljJ and FljL flagellins are divergent from those of the other four flagellins. Since these flagellins are the first to be assembled in the flagellar filament, one or both might have specialized to facilitate the initiation of filament assembly.

Circularity of the Caulobacter crescentus chromosome determined by pulsed-field gel electrophoresis.

Previous genetic analyses of the Caulobacter crescentus chromosome have resulted in the construction of a linear genetic map. To establish the circularity of the C. crescentus chromosome, restriction fragments generated by digestion with AseI and SpeI were analyzed by pulsed-field gel electrophoresis and Southern hybridization. The size of each fragment was calculated and used to demonstrate that C. crescentus has a genome size of approximately 4,000 kilobases. In addition, both enzymes gave rise to large DNA fragments which contained genes from both ends of the genetic map. Thus, there is physical linkage between the genes at the ends of the genetic map and the chromosome is circular. Since this region of the chromosome appears to contain the replication terminus, we propose that recombination occurs at a high frequency in the vicinity of the terminus. This high frequency of recombination would prevent genetic linkage from being observed between genes on opposite sides of the terminus. Additional experiments using insertions which introduced new AseI and DraI restriction sites into the genome allowed us to calculate the physical distance between genes located in the vicinity of the replication terminus.

Use of pulsed field gel electrophoresis and transposon mutagenesis to estimate the minimal number of genes required for motility in Caulobacter crescentus.

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.