Abstract:
:Understanding molecular mechanisms in the context of living cells requires the development of new methods of in vivo biochemical analysis to complement established in vitro biochemistry. A critically important molecular mechanism is genetic recombination, required for the beneficial reassortment of genetic information and for DNA double-strand break repair (DSBR). Central to recombination is the RecA (Rad51) protein that assembles into a spiral filament on DNA and mediates genetic exchange. Here we have developed a method that combines chromatin immunoprecipitation with next-generation sequencing (ChIP-Seq) and mathematical modeling to quantify RecA protein binding during the active repair of a single DSB in the chromosome of Escherichia coli. We have used quantitative genomic analysis to infer the key in vivo molecular parameters governing RecA loading by the helicase/nuclease RecBCD at recombination hot-spots, known as Chi. Our genomic analysis has also revealed that DSBR at the lacZ locus causes a second RecBCD-mediated DSBR event to occur in the terminus region of the chromosome, over 1 Mb away.
journal_name
Proc Natl Acad Sci U S Aauthors
Cockram CA,Filatenkova M,Danos V,El Karoui M,Leach DRdoi
10.1073/pnas.1424269112subject
Has Abstractpub_date
2015-08-25 00:00:00pages
E4735-42issue
34eissn
0027-8424issn
1091-6490pii
1424269112journal_volume
112pub_type
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