Figure 6.

S1 mapping analysis of the 3′-end of RNA transcripts produced by transcriptional arrest and termination. (A) Sequences of the template DNA near the region of triplex site, S1 probe (probe 42), marker 33, and marker 18 are shown. Location of the probe 42 is underlined in the template DNA. Transcriptional arrest and termination sites are shown in two square boxes. Psoralen crosslink site at 185 is indicated by an arrow, and two T residues involved in crosslink formation are highlighted. (B) Analysis of the reaction products of an S1 nuclease mapping experiment. RNA transcripts were hybridized to probe 42 labeled at its 3′-end with ³²P and digested with S1 nuclease, and DNA fragments were analyzed on a sequencing gel. Lane 1, a full-length (340 nucleotides) runoff transcript prepared by using unmodified template. Lane 2, RNA transcripts arising from only termination of elongation were generated by using P_acr probe to form a stable triple helix before performing in vitro transcription reactions. Lane 3, RNA transcripts produced from DNA templates damaged with psoralen adducts. Psoralen probe, P15, was used to form psoralen adducts in the DNA template and in vitro transcription was performed without isolation of unmodified template. Lane M, marker lane containing probe 42, marker 33, and marker 18 oligonucleotides. Sequences of termination and arrest sites are labeled.