Isolation and characterization of four plasmids from Bacillus subtilis

Abstract

Nineteen Bacillus subtilis isolates obtained from type culture collections were examined for the presence of covalently closed circular duplex deoxyribonucleic acid molecules by the technique of cesium chloride-ethidium bromide density gradient centrifugation. Four of the 19 strains tested carried covalently closed circular molecules. Two of these strains (IFO3022, IFO3215) harbored a similar plasmid with a molecular weight of 5.4 x 106. The other two strains (IAM1232, IAM1261) carried 4.9 x 106-and 5.3 x 10fi-dalton plasmids, respectively. These plasmid-harboring strains did not show phenotypic traits such as antibiotic resistance or bacteriocin production. The plasmid deoxyribonucleic acids were digested by three restriction endonucleases, EcoRI, HindIII, and BamNI, and were classified into three different types from their electrophoretic patterns in agarose gels.

Rolling-circle plasmids from Bacillus subtilis: complete nucleotide sequences and analyses of genes of pTA1015, pTA1040, pTA1050 and pTA1060, and comparisons with related plasmids from Gram-positive bacteria

“…The production of trace amounts of ssDNA replication intermediates by each of these plasmids indicates that they all use the rolling-circle mechanism of replication [26]. Plasmids isolated in independent studies by Hara et al [9] (pUH series); Tanaka et al [5,6] (pLS series); Darabi et al [27] (pBS2); Devine et al [28] (pBAA1), and by Poluektova et al [29] (p1410) can also be classi¢ed into one of these six groups. An RCR plasmid belonging to a seventh group, represented by pFTB14, was isolated from the Bacillus amyloliquefaciens strain S294 [8].…”

“…Plasmid size in kbp. pTA series according to Uozumi et al [7]; pLS series according to Tanaka and coworkers [5,6]; pUH series are according to Hara et al [9].…”

Rolling-circle plasmids fromBacillus subtilis: complete nucleotide sequences and analyses of genes of pTA1015, pTA1040, pTA1050 and pTA1060, and comparisons with related plasmids from Gram-positive bacteria

“…The production of trace amounts of ssDNA replication intermediates by each of these plasmids indicates that they all use the rolling‐circle mechanism of replication [26]. Plasmids isolated in independent studies by Hara et al [9](pUH series); Tanaka et al [5, 6](pLS series); Darabi et al [27](pBS2); Devine et al [28](pBAA1), and by Poluektova et al [29](p1410) can also be classified into one of these six groups. An RCR plasmid belonging to a seventh group, represented by pFTB14, was isolated from the Bacillus amyloliquefaciens strain S294 [8].…”

“… b Plasmid size in kbp. pTA series according to Uozumi et al [7]; pLS series according to Tanaka and coworkers [5, 6]; pUH series are according to Hara et al [9]. …”

“…Naturally occurring rolling‐circle replicating (RCR) plasmids form the basis of many cloning and expression vectors for Gram‐positive bacteria. Although the highly transformable Bacillus subtilis strain 168, which has become the paradigm for most studies, does not harbor endogenous plasmids, the existence of B. subtilis plasmids has been known for quite some time [5–11]. Since these plasmids do not confer easily selectable phenotypes to their hosts, most cloning vectors used today for B. subtilis are based on RCR plasmids from other Gram‐positive bacteria, like staphylococci and streptococci [1–3, 12–15].…”