Bacillus subtilis is the best-characterized member of the Gram-positive bacteria. Its genome of 4,214,810 base pairs comprises 4,100 protein-coding genes. Of these protein-coding genes, 53% are represented once, while a quarter of the genome corresponds to several gene families that have been greatly expanded by gene duplication, the largest family containing 77 putative ATP-binding transport proteins. In addition, a large proportion of the genetic capacity is devoted to the utilization of a variety of carbon sources, including many plant-derived molecules. The identification of five signal peptidase genes, as well as several genes for components of the secretion apparatus, is important given the capacity of Bacillus strains to secrete large amounts of industrially important enzymes. Many of the genes are involved in the synthesis of secondary metabolites, including antibiotics, that are more typically associated with Streptomyces species. The genome contains at least ten prophages or remnants of prophages, indicating that bacteriophage infection has played an important evolutionary role in horizontal gene transfer, in particular in the propagation of bacterial pathogenesis.

The approximately 10 kbp region encompassing nprB and argJ at 102" on the Bacillus subtilis chromosome was sequenced, revealing 12 ORFs, four known genes (argJ, argC, ipi and nprB) and two genes, yitV and yitS, whose products respectively display significant homology with L-gulono-y-lactone oxidase of rat and dihydrofolate reductase of Staphy/ococcus aureus. The data also indicated that nprB mapped to a different position than previously published.Keywords : Bacillus subtilis, genome sequencing, 102' region, argJ, nprB Within the European programme for sequencing the Bacillus subtilis genome, a 60 kbp DNA fragment located between nprB (95") and arg] (102") was originally assigned to our group. Since, as described in this paper, nprB was, in fact, found to be located approximately 8 kb from arg], the assignment was redefined and the region to be sequenced was extended from arg] (102") to addAB (89"). In this first communication, we describe our cloning and sequencing strategy and we report the sequencing and gene features of the 10336 bp region from nprB to arg].For this study, we employed a chromosome walking strategy using the integrative plasmid pDIA5304 (Glaser et al., 1993), into which we inserted a 1 kb fragment from arg] obtained from pUL710. This allowed us, after digestion of the chromosomal DNA with an appropriate restriction enzyme ( S a d ) , to pick up a fragment of more than 10 kbp adjacent to argJ. The recombinant plasmid, pOMG3001, was propagated in Escherichia coli TP611, a mutant strain (pcnB) which facilitates cloning of B. subtilis DNA by reducing the copy number of ColEl derivatives (Xu et al., 1993). The integrity of the clone was verified by Southern hybridization. For shotgun analysis, pOMG3001 was subjected to a nebulization procedure and 1-1-5 kb fragments were subcloned into pUCl8. After identification and selection by Southern hybridization, the fragments corresponding to the insert were sequenced. The sequencing reactions were performed using the Taq Dye Primer Cycle Sequencing kit and the Taq Dye Deoxy Terminator Cycle Sequencing kit, followed by analysis with a 373A DNA sequencer Abbreviation: SD, Shine-Dalgarno.The EMBL accession number for the nucleotide sequence reported in this paper is 279580.from Applied Biosystems. The sequences were compiled using the Autoassembler program (Perkin Elmer) and analysed for possible ORFs (GENEMARK). The predicted amino acid sequence of the products of each ORF was used to search for similarity to sequences reported previously in databases using BLAST/FASTA.From the sequence data, all six possible reading frames were screened with the DNA Strider program. Based on starting codons ATG, GTG or TTG preceded by a putative Shine-Dalgarno (SD) sequence, 12 putative ORFs were detected. The direction of transcription and translation of all except orf2 ( yitS) and orf. ( yitU), was, like the majority of genes on the B. subtilis chromosome, 0