Protein affinity maturation in vivo using E-coli mutator cells

This protocol provides a simple in vivo strategy for introducing random mut ations to a target DNA sequences using E. coli mutator cells. The method ha s been used in our laboratory for affinity maturation of proteins encoded b y target DNA sequences. Selection conditions can be modified for antibody f ragments with increased production levels. Growth conditions in E. coli mut ator cells can be adjusted to introduce a single random point mutation per kilobase of DNA, approximately equivalent to one codon change per scFv frag ment. (C) 2001 Elsevier Science B.V. All rights reserved.