Because of the potential use of the Bacilli for genetic manipulation, experiments were undertaken to investigate the usefulness of Bacillus subtilis and Bacillus licheniformis strain LO2, as hosts. Attempts at shotgun-cloning directly in B. subtilis met with repeated failure. However, subsequently the Φ3T-thyP3 gene, pC194 chloramphenicol acetyl transferase gene, B. licheniformis 749/C penP gene and the E. coli lacZ gene were expressed in B. subtilis when cloned into the plasmid vector pAB224 or one of its derivatives. Consequently, such plasmids are useful vectors for genetic manipulation in B. subtilis. The properties of the thyP3-containing hybrids were investigated. Of particular interest was the finding that monomeric plasmid DNA, containing the thyP3 gene, was active in the transformation of competent B. subtilis cells. Transformation resulted in integration of the thyP3-containing region of the plasmid into the host chromosome. The secretion of fusion proteins by B. subtilis was investigated employing the B. licheniformis 749/C penicillinase protein signal-peptide. This resulted in the secretion of the E. coli β-galactosidase enzyme from the B. subtilis cell. The thermotolerant Bacillus licheniformis strain LO2 was investigated as a possible host for genetic manipulation. A series of mutant strains were isolated but the induction of competence in two such strains could not be achieved. Additionally, transformation of protoplasts of strain LO2 could not be demonstrated. Thus contrary to previous hopes, at present this strain does not appear to be suitable as a host for genetic manipulation.
| Identifer | oai:union.ndltd.org:bl.uk/oai:ethos.bl.uk:238004 |
| Date | January 1983 |
| Creators | Barstow, David A. |
| Publisher | University of Warwick |
| Source Sets | Ethos UK |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Source | http://wrap.warwick.ac.uk/102341/ |
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