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. 2014 Jul 29;111(32):E3269–E3276. doi: 10.1073/pnas.1406234111

Fig. 3.

Fig. 3.

Transcriptional efficiency of RNA pol II is greatly reduced by the linkage alteration in the DNA template. (A) Scaffold of RNA, template DNA, and nontemplate DNA for runoff elongation through the 2′–5′ linkage site in the template. The red star refers to the 2′–5′ linkage in the DNA. (B) RNA pol II transcription products of 3′–5′–linked (left part of the gel) and 2′–5′–linked (right part of the gel) scaffolds in the presence of 25 μM NTP. Time points are 30 s, 2 min, 5 min, 20 min, and 1 h from left to right. Numbers on the left refer to lengths of RNA elongation products; positions of these RNA products on the DNA template are partly shown on the right. The boxed area shows the major difference with linkage alteration. (C) Scaffolds used to study every transcriptionally slowed down step. Three primers refer to three stalled positions in the DNA template. The red star refers to the 2′–5′ linkage. (D) Catalytic constants (kpol) and specificity constants (kpol/Kd,app) of pol II transcription through 3′–5′ linkage (blue) and 2′–5′ linkage (red).