ChIP-chip to analyze the binding of replication proteins to chromatin using oligonucleotide DNA microarrays.
Chromatin immunoprecipitation (ChIP) is a widely used method to study the interactions between proteins and discrete chromosomal loci in vivo. Originally, ChIP was developed for analysis of protein associations with DNA sequences known or suspected to bind the protein of interest. The advent of DNA microarrays has enabled the identification of all DNA sequences enriched by ChIP, providing a genomic view of protein binding. This powerful approach, termed ChIP-chip, is broadly applicable and has been particularly valuable in DNA replication studies to map replication origins in Saccharomyces cerevisiae based on the association of replication proteins with these chromosomal elements. We present a detailed ChIP-chip protocol for S. cerevisiae that uses oligonucleotide DNA microarrays printed on polylysine-coated glass slides and can also be easily adapted for commercially available high-density tiling microarrays from NimbleGen. We also outline general protocols for data analysis; however, microarray data analyses usually must be tailored specifically for individual studies, depending on experimental design, microarray format, and data quality.
Viggiani
Christopher J
CJ
Molecular and Computational Biology Program, University of Southern California, Los Angeles, CA, USA.
Aparicio
Jennifer G
JG
Aparicio
Oscar M
OM
D002843:Chromatin / Q000378:metabolism*; D047369:Chromatin Immunoprecipitation / Q000379:methods* / Q000706:statistics & numerical data; D003432:Cross-Linking Reagents; D004261:DNA Replication; D004271:DNA, Fungal / Q000096:biosynthesis / Q000302:isolation & purification; D004268:DNA-Binding Proteins / Q000378:metabolism*; D003627:Data Interpretation, Statistical; D005456:Fluorescent Dyes; D009693:Nucleic Acid Hybridization; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods* / Q000706:statistics & numerical data; D018741:Replication Origin; D012441:Saccharomyces cerevisiae / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism
D002843:Chromatin; D003432:Cross-Linking Reagents; D004271:DNA, Fungal; D004268:DNA-Binding Proteins; D005456:Fluorescent Dyes; D029701:Saccharomyces cerevisiae Proteins