1887

Abstract

Genome sequencing is making a profound impact on microbiology. Currently, however, only one plant pathogen genome sequence is publicly available and no genome-sequencing project has been initiated for any species of the genus , which includes several important plant pathogens. This paper describes a targeted sample sequencing approach to study the genome of subsp. (), a major soft-rot pathogen of potato. A large insert DNA (approx. 115 kb) library of was constructed using a bacterial artificial chromosome (BAC) vector. Hybridization and end-sequence data revealed two overlapping BAC clones that span an entire gene cluster. Random subcloning and one-fold sequence coverage (>200 kb) across these BACs identified 25 (89%) of 28 genes predicted from the orthologous cluster of . Regions flanking the cluster contained orthologues of known or putative pathogenicity operons from other species, including (), and (), sequences similar to genes from the plant pathogen , including haemagglutinin-like genes, and sequences similar to genes involved in rhizobacterium–plant interactions. Approximately 10% of the sequences showed strongest nucleotide similarities to genes in the closely related model bacterium and animal pathogen . However, the positions of some of these genes were different in the two genomes. Approximately 30% of sequences showed no significant similarity to any database entries. A physical map was made across the genomic region spanning the cluster by hybridization to the BAC library and to digested BAC clones, and by PCR between sequence contigs. A multiple genome coverage BAC library and one-fold sample sequencing are an effective combination for extracting useful information from important regions of the genome, providing a wealth of candidate novel pathogenicity genes for functional analyses. Other genomic regions could be similarly targeted.

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2002-05-01
2020-04-08
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