Description

This track displays validation data from ChIP-chip experiments on four factors in IMR90 cells using a condensed array covering putative TAF1 binding sites. This track may be used to validate the whole genome scan shown in the LI/Ng TAF1 IMR90 track (Ludwig Institute, UCSD ChIP-chip (TAF1) genome scan). All four factors — Pol2, H3ac, H3K4me2, and TAF1 itself — are associated with the start of transcribed genes. Thus, there should be a very strong correlation between the signals shown in this track and the LI/NG TAF1 IMR90 track.

TAF1 is a component of TFIID, which is itself a component of the pre-initiation complex that assembles on promoter regions. Pol2, more fully known as RNA Polymerase II, is the enzyme responsible for transcription of mRNA. The specific antibody against Pol2, 8WG16 from the Abcom catalog, binds specifically to the non-phosphorylated form of Pol2 which is associated with the pre-initiation complex. H3ac and H3K4me2 are forms of histone H3 that are associated with transcriptionally-active chromatin.

Methods

For the whole genome scan, chromatin from IMR90 cells lines was cross-linked, precipitated with TAF1 antibody (sc-735, Santa Cruz), sheared, amplified and hybridized to 38 high-density oligonucleotide arrays (NimbleGen). These arrays contain a total of 14,535,659 50-mer oligonucleotides, positioned at every 100 base pairs throughout the human genome (UCSC hg16). Using this set of arrays, a total of 9,966 clusters of TAF1 binding sites were identified. The whole genome scan data can be viewed in the LI/Ng TAF1 IMR90 track (Ludwig Institute, UCSD ChIP-chip (TAF1) genome scan).

To verify the binding of TAF1 to these sequences, a condensed array was designed containing a total of 379,521 oligonucleotides to represent the 9,966 putative TAF1 binding sequences plus 29 control genomic loci at 100 bp resolution. Using these condensed arrays, two independent chromatin immunoprecipitation (ChIP) experiments were performed with the antibodies against TAF1, Pol2, acetylated histone 3 and dimethylated K4 histone 3. A total of 8,597 TAF1 binding regions, ranging in size from 400 bp to 9.8 Kbp, were confirmed by the TAF1 replicate experiments.

The raw data have been deposited in GEO (GSE2672) and will be released following publication of the paper.

Verification

The peaks from genome scan experiments were verified using condensed arrays, as described in the Methods section.

References

Kim, T.H., Barrera, L.O., Zheng, M., Qu, C., Singer, M.A., Richmond, T.A., Wu, Y., Green, R.D. and Ren, B. A high-resolution map of active promoters in the human genome. Nature 436, 876-880 (2005).