Intact recombineering of highly repetitive DNA requires reduced induction of recombination enzymes and improved host viability

Recombineering technology permits flexible engineering of large DNA in Escherichia coli without dependence on suitably placed restriction sites. However, recombineering is limited for modifying highly repetitive DNA because of its potential to trigger instability by uncontrolled self-recombination of the repeats. In this study, induction of the recombineering enzymes and growth condition of the host are optimized to demonstrate intact modification of bacterial artificial chromosomes (BACs) containing long arrays of centromeric alpha satellite repeats. This optimized recombineering protocol may be useful for manipulation of other biologically important repetitive DNAs, including trinucleotide repeat expansions and homologous gene families, to facilitate their functional studies. (c) 2008 Elsevier Inc. All rights reserved.