RNA polymerases are highly regulated molecular machines. We present a method 
(global run-on sequencing, GRO-seq) that maps the position, amount, and 
orientation of transcriptionally engaged RNA polymerases genome-wide. In this 
method, nuclear run-on RNA molecules are subjected to large-scale parallel 
sequencing and mapped to the genome. We show that peaks of promoter-proximal 
polymerase reside on approximately 30% of human genes, transcription extends 
beyond pre-messenger RNA 3' cleavage, and antisense transcription is prevalent. 
Additionally, most promoters have an engaged polymerase upstream and in an 
orientation opposite to the annotated gene. This divergent polymerase is 
associated with active genes but does not elongate effectively beyond the 
promoter. These results imply that the interplay between polymerases and 
regulators over broad promoter regions dictates the orientation and efficiency 
of productive transcription.