A STABLE AND EFFICIENT TRANSFORMATION SYSTEM FOR BUTYRIVIBRIO-FIBRISOLVENS OB156

A 9.5-kb shuttle vector capable of replication and selection in both E scherichia coli and Butyrivibrio fibrisolvens was constructed. Plasmid pUC118 provided replication functions and ampicillin resistance selec tion in E. coli. In B. fibrisolvens, replication was controlled by the native plasmid pRJF1 from strain OB156, and selectability was provide d by a 3.5-kb fragment of plasmid pAM beta 1 containing the erythromyc in resistance gene. Optimum conditions for transformation were 15 kV/c m, 2 h recovery, and plating in an agar overlay on medium containing 1 0 mu g erythromycin/ ml. Maximum efficiency was 1.1 x 10(5) transforma nts per mu g plasmid DNA (average 3 x 10(4)), and restriction mechanis ms reduced efficiency by a factor of 2 x 10(2). Nonselective growth fo r 200 generations gave no measurable loss of plasmid.