Rhodobacter capsulatus

In the collaborative effort of the laboratory of Robert Haselkorn at the University of Chicago and the laboratory of Vaclav Paces at the Institute of Molecular Genetics, Academy of Sciences of the Czech Republic, the genome of Rhodobacter capsulatus has been completely sequenced.

Rhodobacter capsulatus is a purple, nonsulfur photosynthetic bacterium. The biochemical versatility to choose between photo- and heterotrophic growth and nitrogen fixation is packed in a genome that is 75% of the size of the Escherichia coli genome. Ease of plating and generation of mutations together with convenient systems for cloning and genetic analysis make R. capsulatus a popular model system for studies of photochemical reaction centers and nitrogen fixation. The GTA (gene transfer agent) has been used for insertional mutagenesis, gene replacement, and linkage analysis. The development of shuttle vectors based on broad host range plasmids made possible efficient random and site-specific mutagenesis with transposons and gene cloning by mutant complementation.

The R. capsulatus genome consists of a single chromosome containing 3.5 Mb and a circular plasmid of 134 kb. The genome sequencing started in 1992. It began with a fine-structure restriction map of an overlapping set of cosmids that covered the genome. Cosmid sequencing led to a gapped genome that was filled by primer walking on the chromosome and by using lambda clones. Methods had to be developed to handle strong stops in the high GC (68%) inserts. Annotation was done at the Institute of Molecular Genetics and with the ERGO system at Integrated Genomics. From these data the cell's metabolism was reconstructed. It was possible to recognize 3709 orfs of which functional assignments could be made with high confidence to 2392 (65%). Unusual features include the presence of numerous cryptic phage genomes embedded in the chromosome.

This project was supported by grants No XXXX to Robert Haselkorn at the University of Chicago a by grants xxxx to Vaclav Paces at the Institute of Molecular Genetics in Prague. In the middle of the project, when we started its most demnanding part, e.i. closing the gaps and putting the large contigs into one contiguous DNA sequence, funding was not renewed, and the project was stopped for four years. It has been finished in 2006.