Chromatin immunoprecipitation to investigate protein-DNA interactions during genetic recombination.
Chromatin immunoprecipitation is a technique that allows one to examine the in vivo localization of proteins to DNA. This technique is well suited for studying genetic recombination since it can provide both a temporal and spatial assessment of the dynamic association of proteins with DNA in both wild-type and mutant backgrounds. To perform this procedure, cells undergoing a synchronous recombination event are treated with a crosslinking agent. Following cell lysis and shearing of the DNA, immunoprecipitation is used to isolate the protein of interest, along with any DNA that is crosslinked to the protein. Polymerase chain reaction (PCR) is then used to determine the relative amounts of DNA associated with the protein of interest throughout the recombination event. This in vivo chemical crosslinking technique can be used to localize proteins to both double-strand breaks and recombination intermediates.
Goldfarb
Tamara
T
Department of Molecular Biology and Genetics, Cornell University, Ithaca, NY, USA.
Alani
Eric
E
D002843:Chromatin / Q000235:genetics* / Q000302:isolation & purification; D015825:Chromosomes, Fungal / Q000235:genetics; D003433:Crosses, Genetic; D004271:DNA, Fungal / Q000235:genetics / Q000302:isolation & purification; D007202:Indicators and Reagents; D008957:Models, Genetic; D016133:Polymerase Chain Reaction / Q000379:methods; D011995:Recombination, Genetic; D012441:Saccharomyces cerevisiae / Q000235:genetics*
D002843:Chromatin; D004271:DNA, Fungal; D007202:Indicators and Reagents