A dual gene-specific mutator system installs all transition mutations at similar frequencies in vivo.

Targeted in vivo hypermutation accelerates directed evolution of proteins through concurrent DNA diversification and selection. Although systems employing a fusion protein of a nucleobase deaminase and T7 RNA polymerase present gene-specific targeting, their mutational spectra have been limited to exclusive or dominant C:G→T:A mutations. Here we describe eMutaT7transition, a new gene-specific hypermutation system, that installs all transition mutations (C:G→T:A and A:T→G:C) at comparable frequencies.