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Quantification of Jn signal end breaks in developing B cells by blunt-end linker ligation and qPCR

By John D. Curry, Lydia Li and Mark S. Schlissel

Abstract Introduction of a double-strand DNA break at the junction between a rearranging gene segment and its flanking recombination signal sequence (RSS) is the first step of V(D)J recombination. Such DNA breaks can be detected by either Southern blot hybridization or ligation-mediated PCR. While Southern blotting is easily quantifiable, it is often insufficiently sensitive and while LM-PCR is far more sensitive, it is poorly quantifiable. Reported here is a LM-qPCR assay which relies on real-time qPCR to provide an absolute measure of recombinase-mediated, or any other specific, double-strand DNA break i

Year: 2004
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