Figure 4

HQ formation upstream of a T7 promoter in plasmid detected by RNA polymerase arrest. (a) The plasmid was first transcribed with T7 RNAP to induce the formation of HQ between the SP6 and T7 promoter. Then the plasmid was transcribed by SP6 RNAP to produce fluorescently labeled RNA. The T7 RNAP was captured with an excess amount of competitive DNA to prevent further T7 transcription. SP6 transcripts were resolved by denaturing polyacrylamide gel electrophoresis. (b) HQ formation between the SP6 and T7 promoter caused premature termination of SP6 transcription (blue arrowhead). FL, Full-length transcript. PT, prematurely terminated transcript. The two bands in lane 1 are the marker transcripts of the two linearized plasmids, approximately representing the size of the prematurely terminated transcripts and the full-length transcripts. The bottom marker was shorter than the distance between the SP6 promoter and PHQS by 19 nucleotides.