Transformation of vegetative cells of Bacillus anthracis with plasmid DNA

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 103 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 104 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.