Improved genome-wide localization by ChIP-chip using double-round T7 RNA polymerase-based amplification

Chromatin immunoprecipitation combined with DNA microarrays (ChIP-chip) is a powerful technique to detect in vivo protein-DNA interactions. Due to low yields, ChIP assays of transcription factors gener- ally require amplification of immunoprecipitated genomic DNA. Here, we present an adapted linear amplification method that involves two rounds of T7 RNA polymerase amplification (double-T7). Using this we could successfully amplify as little as 0.4ng of ChIP DNA to sufficient amounts for microarray analysis. In addition, we compared the double-T7 method to the ligation-mediated polymerase chain reaction (LM-PCR) method in a ChIP-chip of the yeast transcription factor Gsm1p. The double-T7 protocol showed lower noise levels and stronger binding signals compared to LM-PCR. Both LM-PCR and double-T7 identified strongly bound genomic regions, but the double-T7 method increased sensi- tivity and specificity to allow detection of weaker binding sites.