Abstract: This disclosure provides methods and compositions for targeted mutagenesis of specific genes in a bacterial strain. By inducibly over-expressing error-prone polymerases such as Pol IV or Pol V in conjunction with nickase in a bacterial strain, and housing the targeted gene(s) on an episome or plasmid which contains one or more nickase recognition sequences, the targeted gene(s) can be selectively mutated at rates significantly greater than genes contained on the chromosome. The methods disclosed herein are useful for engineering desirable bacterial phenotypes and novel strains, including for example strains useful for treating or degrading waste and/or environmental contaminants, for optimizing bioprocesses, and for converting low-value feed-stock into value-added products.

Abstract: This disclosure provides methods and compositions for targeted mutagenesis of specific genes in a bacterial strain. By inducibly over-expressing error-prone polymerases such as Pol IV or Pol V in conjunction with nickase in a bacterial strain, and housing the targeted gene(s) on an episome or plasmid which contains one or more nickase recognition sequences, the targeted gene(s) can be selectively mutated at rates significantly greater than genes contained on the chromosome. The methods disclosed herein are useful for engineering desirable bacterial phenotypes and novel strains, including for example strains useful for treating or degrading waste and/or environmental contaminants, for optimizing bioprocesses, and for converting low-value feed-stock into value-added products.