Transformation of vegetative cells of Bacillus anthracis with plasmid DNA

1 Division of Biologics, Centre for Applied Microbiology and Research, Porton Down, Salisbury, Wiltshire SP4 0JG, UK 2 School of Pure and Applied Biology, University of Wales College of Cardiff, PO Box 915, Cardiff CF1 3TL, UK ABSTRACT Summary: Methods have been developed for chemical transformation...

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Published inJournal of general microbiology Vol. 136; no. 7; pp. 1211 - 1215
Main Authors Quinn, Conrad P, Dancer, Brian N
Format Journal Article
LanguageEnglish
Published England Soc General Microbiol 01.07.1990
Subjects
Bacillus anthracis
Bacillus anthracis - genetics
Bacillus anthracis - growth & development
Buffers
Culture Media
Electricity
Hydrogen-Ion Concentration
Plasmids
Polyethylene Glycols
Transformation, Bacterial
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Summary:1 Division of Biologics, Centre for Applied Microbiology and Research, Porton Down, Salisbury, Wiltshire SP4 0JG, UK 2 School of Pure and Applied Biology, University of Wales College of Cardiff, PO Box 915, Cardiff CF1 3TL, UK ABSTRACT Summary: Methods have been developed for chemical transformation and electro-transformation (electroporation) of vegetative cells of Bacillus anthracis with supercoiled plasmid DNA. Chemical transformation was dependent on incubation in Tris/HCl with osmotic support and transformation with plasmid DNA was effected by treatment with polyethylene glycol 3350. Maximum transformation frequencies were 3.8 x 10 –5 transformant c.f.u. per viable c.f.u. (1 x 10 3 c.f.u. per µg DNA). Optimal frequencies were pH dependent and were affected by growth-medium composition. Transformation was not observed with linear or multimeric plasmid DNA. Electro-transformation of B. anthracis using high field intensity electroporation was dependent on the composition of both the growth medium and the electroporation buffer. Maximum electro-transformation frequencies were 5.3 x 10 –4 c.f.u. per viable c.f.u. (2.6 x 10 4 c.f.u. per µg DNA). The use of early exponential phase cells was critical to both procedures and the maximum efficiency (c.f.u. per µg DNA) of each system was strain dependent under the conditions described. Present address: Laboratory of Microbial Ecology, 30–309 NIDR, National Institutes of Health, Bethesda, Maryland 20892-0030, USA.
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ISSN:0022-1287
DOI:10.1099/00221287-136-7-1211