New Chromatin Run-On Reaction Enables Global Mapping of ActiveRNA Polymerase Locations in an Enrichment-free Manner br

The development of a simple and cost-effectivemethod to map the distribution of RNA polymerase II (RNPII)genome-wide at a high resolution is highly beneficial to studycellular transcriptional activity. Here we report a mutation-basedand enrichment-free global chromatin run-on sequencing (mGRO-seq) technique to locate active RNPII sites genome-wide at near-base resolution. An adenosine triphosphate (ATP) analog namedN6-allyladenosine triphosphate (a6ATP) was designed and couldbe incorporated into nascent RNAs at RNPII-located positionsduring a chromatin run-on reaction. By treatment of the run-onRNAs with a mild iodination reaction and subjection of theproducts to reverse transcription into complementary DNA(cDNA), base mismatch occurs at the original a6A incorporationsites, thus making the RNPII locations detected in the high-throughput cDNA sequencing. The mGRO-seq yields both the map ofRNPII sites and the chromatin RNA abundance and holds great promise for the study of single-cell transcriptional activity.