Figure 8.

RNA polymerase II associates with both alleles in a monoallelically expressing cell line. (A) Recruitment of RNA polymerase II to the IGF2BP1 promoter was examined by ChIP in monoallelically expressing GM7007 cells. DNA recovered from chromatin that had been immunoprecipitated with anti-RNA polymerase II antibodies (Pol2) was amplified and sequenced for allelic association. Sequencing results (bottom) reveal that both alleles of the monoallelically expressing cell line GM7007 associate with RNA polymerase II near SNP site rs4794017. In contrast, sequencing of DNA from "no antibody" ChIP reactions failed to produce sequence reads. (B) Allele specificity of precursor mRNA was determined by sequencing of cDNA prepared from total RNA of GM7007 cells. RNA had been extensively pretreated with DNase I to eliminate gDNA prior to reverse transcription by RT. Subsequently, cDNA samples were amplified using primers flanking rs4794017. In the absence of RT (-RT), no amplification products were oberved. +RT amplicons were gel-purified and sequenced. Bottom: Sequence traces at the heterozygous SNP site rs4794017 located 1 kb downstream of the transcription initiation site in cDNA of GM7007 indicate a single allele.