Genomewide identification of protein binding locations using chromatin immunoprecipitation coupled with microarray.
Interactions between cis-acting elements and proteins play a key role in transcriptional regulation of all known organisms. To better understand these interactions, researchers developed a method that couples chromatin immunoprecipitation with microarrays (also known as ChIP-chip), which is capable of providing a whole-genome map of protein-DNA interactions. This versatile and high-throughput strategy is initiated by formaldehyde-mediated cross-linking of DNA and proteins, followed by cell lysis, DNA fragmentation, and immunopurification. The immunoprecipitated DNA fragments are then purified from the proteins by reverse-cross-linking followed by amplification, labeling, and hybridization to a whole-genome tiling microarray against a reference sample. The enriched signals obtained from the microarray then are normalized by the reference sample and used to generate the whole-genome map of protein-DNA interactions. The protocol described here has been used for discovering the genomewide distribution of RNA polymerase and several transcription factors of Escherichia coli.
Cho
Byung-Kwan
BK
Department of Bioengineering, University of California-San Diego, La Jolla, CA, USA.
Knight
Eric M
EM
Palsson
Bernhard Ø
BØ
D001483:Base Sequence; D047369:Chromatin Immunoprecipitation; D017931:DNA Primers; D016678:Genome; D020411:Oligonucleotide Array Sequence Analysis; D011485:Protein Binding
D017931:DNA Primers