Bidirectionally transcribed loci were identified in the same way as with pooled FANTOM5 CAGE libraries (see ‘Identification of bidirectionally transcribed loci’ and ‘Expression quantification of bidirectionally transcribed loci and prediction of enhancers’ above) from tag clusters (as defined in 51) derived from pooled HeLa CAGE (mock treated control, hMTR4) libraries. From 5,892 bidirectional loci distant to TSSs and exons, 4196 were predicted to be enhancers based on balanced directionality of transcription of which 3,896 had significantly greater expression than random genomic regions in at least one library. These were then overlapped with the whole set of FANTOM5 CAGE enhancers to estimate the fraction of unseen transcribed enhancers.
