Figure 4.

RNA·DNA hybrids are associated with transcription arrest on TTC templates in vitro. Pairs of transcription reactions were done in the absence (−) or presence (+) of RNase H on supercoiled templates containing the triplet repeat sequences (CTG·CAG)₈₈ (lanes 1 and 2), (GAA·TTC)₈₈ (lanes 3 and 4) and (TTC·GAA)₈₈ (lanes 5 and 6). The gel image shows the products of those reactions. To the right of the gel image the length in bases of select DNA size markers are indicated. At the far right of the figure is a scan of lanes 3 (dark line) and 4 (gray line) which highlights the RNase H mediated shift in truncation points from promoter distal (arrowhead) to promoter proximal within the GAA repeat. All the templates contain the sequence for a self-cleaving ribozyme that cuts the transcript 270 bases 3′ to the end of the repeat tract producing a full-length transcript of 590 bases. Transcription from the phage T7 promoter started 55 bases 5′ to the GAA·TTC insert. Transcripts were end-labeled by including gamma ³²P-GTP in the reaction.