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Title: Recombination at a 246 bp interrupted palindrome in Escherichia coli
Author: Eykelenboom, John K.
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 2006
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Long DNA palindromes are sites of genome instability (deletions, gene amplification and translocations) in eukaryotic cells. In both prokaryotic and eukaryotic cells they are sites of DNA breakage that can be repaired by homologous recombination. Genetic evidence suggests that in E. coli breakage is mediated by the SbcCD complex (Rad50/Mre11) that can cleave DNA-hairpin structures. Here an arabinose inducible-sbcDC (pBAD-sbcDC) system has been constructed and used to obtain genetic and in vivo physical evidence of DNA double-strand breaks (DSBs) at a 246 bp interrupted palindrome. These breaks are shown to have two ends suggesting that they occur behind the replication fork probably on the lagging strand. A two-ended break is not compatible with the palindrome blocking DNA replication and cleavage causing the fork to collapse. Repair of the breaks requires homologous recombination facilitated by the RecA and RecBCD proteins as well as the Holliday junction resolves RuvABC and RecG. Contrary to previous reports this study rules out a significant role of the RecFOR proteins in the repair of breaks and implicates a role for the PriA protein – probably to establish replication forks at the break site. Removing the XerCD/dif system (chromosome dimer resolution) has a minor negative effect on cell viability suggesting that chromosome crossover events occur at a detectable (but unquantifiable) frequency. There is also a requirement for cleavable LexA protein demonstrating that the SOS response is required for efficient repair.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available