Rational design of a plasmid origin that replicates efficiently in both gram-positive and gram-negative bacteria.

Abstract

Most plasmids replicate only within a particular genus or family.Here we describe an engineered high copy number expression vector, pBAV1K-T5, that produces varying quantities of active reporter proteins in Escherichia coli, Acinetobacter baylyi ADP1, Agrobacterium tumefaciens, (all gram-negative), Streptococcus pneumoniae, Leifsonia shinshuensis, Peanibacillus sp. S18-36 and Bacillus subtilis (gram-positive).Our results demonstrate the efficiency of pBAV1K-T5 replication in different bacterial species, thereby facilitating the study of proteins that don't fold well in E. coli and pathogens not amenable to existing genetic tools.