Figure 3

TFIIB and TFIID bind internal positions in RefSeq genes and are associated with internally initiated transcripts. (A) Real-time PCR quantitation of DNA fragments precipitated in a ChIP assay using 3 μg TFIIB antibody (black bars), 10 μl of TFIID antibody (gray bars) or 3 μg Gal4 antibody as a control IgG (white bars). Regions interrogated correspond to a subset of genes with internal sites that were chosen at random. The levels of Ins1 and CBP 5' promoters precipitated are included as controls. Negative regions included the repressed HBB and IL2 genes. Data is presented as percent input and error is SEM. 5', internal, and 3' refer to the positions of the TFIIB GST relative to RefSeq gene boundaries. (B) An agarose gel of the amplified transcript from an internally initiated CBP transcript identified by a modified cap trapping procedure (see methods). Two independent PCR clones (lanes 1 and 2) mapped the start site to nucleotide 11687697 on chromosome 10.