A partial physical map for the chromosome of alkalophilic Bacillus sp. strain C-125

Abstract

Bacillus sp. strain C-125 has been chosen as a model alkalophilic bacterium to understand how adaptation to growth at high pH is achieved. To aid genetic analysis, we have started characterization of its genome. By using the two infrequently-cutting restriction endonucleases, A scI and Sse83871, in conjunction with pulsed-field electrophoretic techniques, the size of the genome was found to be 3.7 Mb. Southern blot analysis of single, double and partial digests of Bacillus sp. strain C-125 DNA, using AscI-linking clones, gene probes and purified Bacillus sp. strain C-125 restriction fragments, allowed a putative chromosome map to be constructed.

Genome Analysis of Facultatively Alkaliphilic Bacillus halodurans C-125

“…NotI (S'-GC/GGCCGCC- 3') did not digest the C-125 chromosome as described in a previous paper (Southerland et al 1993). The cells from 500 111 of culture were harvested by centrifugation, washed once in TE buffer (10 mM Tris-HCI pH 8.0, I mM EDTA), resuspended in the same volume of TE buffer (50 o q, and mixed with 500 111 of 2% Pulsed Field Certified (PFq agarose prewarmed to 50°C.…”

“…The blocks were washed once in TE buffer and then incubated in TE buffer containing I mM phenylrnethylsulfonyl fluoride (PMSF) for I h at room temperature. It appeared that the genome size had been underestimated previously (Southerland 1993) for the reason described above. Restriction endonuc1eases that recognize an 8-bp sequence were tested for their ability to digest the chromosome of strain C-125.…”

“…We report a revision of the genome size from 3.7 Mb (Southerland et al 1993) to 4.25 Mb and a complete AscII Sse8387I physical map of the chromosome of Bacillus halodurans C-125 (Takami et al 1999b We report a revision of the genome size from 3.7 Mb (Southerland et al 1993) to 4.25 Mb and a complete AscII Sse8387I physical map of the chromosome of Bacillus halodurans C-125 (Takami et al 1999b …”