Comparison of sample preparation methods for ChIP-chip assays.
A single chromatin immunoprecipitation (ChIP) sample does not provide enough DNA for hybridization to a genomic tiling array. A commonly used technique for amplifying the DNA obtained from ChIP assays is ligation-mediated PCR (LM-PCR). However; using this amplification method, we could not identify Oct4 binding sites on genomic tiling arrays representing 1% of the human genome (ENCODE arrays). In contrast, hybridization of a pool of 10 ChIP samples to the arrays produced reproducible binding patterns and low background signals. However the pooling method would greatly increase the number of ChIP reactions needed to analyze the entire human genome. Therefore, we have adapted the GenomePlex whole genome amplification (WGA) method for use in ChIP-chip assays; detailed ChIP and amplification protocols used for these analyses are provided as supplementary material. When applied to ENCODE arrays, the products prepared using this new method resulted in an Oct4 binding pattern similar to that from the pooled Oct4 ChIP samples. Importantly, the signal-to-noise ratio using the GenomePlex WGA method is superior to the LM-PCR amplification method.
O'Geen
Henriette
H
University of California-Davis, Davis, CA 95616, USA.
Nicolet
Charles M
CM
Blahnik
Kim
K
Green
Roland
R
Farnham
Peggy J
PJ
D000818:Animals; D001665:Binding Sites; D002478:Cells, Cultured; D047369:Chromatin Immunoprecipitation / Q000379:methods*; D015894:Genome, Human; D006801:Humans; D051379:Mice; D021141:Nucleic Acid Amplification Techniques; D050814:Octamer Transcription Factor-3 / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D016133:Polymerase Chain Reaction
D050814:Octamer Transcription Factor-3