Recombinogenic engineering methodology, also known as recombineering, utilizes 
homologous recombination to create targeted changes in cellular DNA with great 
specificity and flexibility. In Escherichia coli, the Red recombination system 
from bacteriophage lambda has been used successfully to modify both plasmid and 
chromosomal DNA in a highly efficient manner, using either a linear 
double-stranded DNA fragment or a synthetic single-stranded oligonucleotide 
(SSO). The current model for Red/SSO-mediated recombination involves the SSO 
first annealing to a transient, single-stranded region of DNA before being 
incorporated into the chromosome or plasmid target. It has been observed 
previously, in both eukaryotes and prokaryotes, that mutations in the two 
strands of the DNA double helix are 'corrected' by complementary SSOs with 
differing efficiencies. Here we investigate further the factors that influence 
the strand bias as well as the overall efficiency of Red/SSO-mediated 
recombination in E.coli. We show that the direction of DNA replication and the 
nature of the SSO-encoded mismatch are the main factors dictating the 
recombinational strand bias. However, the influence that the SSO-encoded 
mismatch exerts upon the recombinational strand bias is abolished in E.coli 
strains that are defective in mismatch repair (MMR). This reflects the fact that 
different base-base mispairs are corrected by the mutS/H/L-dependent MMR pathway 
with differing efficiencies. Furthermore, our data indicate that transcription 
has negligible influence on the strand bias. These results demonstrate for the 
first time that the interplay between DNA replication and MMR has a major effect 
on the efficiency and strand bias of Red/SSO-mediated recombination in E.coli.