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Schematic of LORAX-seq. Chromatin was isolated from fractionated nuclei using urea and Empigen before digestion with DNase. Transcription complexes were immunoprecipitated, washed, and then treated with phosphatase to remove contaminating 5′ phosphates. Purified transcription complexes were then incubated with purified TFIISin vitroto cleave and elute backtracked RNA, generating new 5′ phosphates for ligation. The elution was purified and prepared for next generation sequencing. Sequencing reads directly correspond with backtracked RNA, providing the precise location that backtracking begins and ends.*denotes steps adapted from POINT.

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