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. 2015 Feb 26;43(5):2625–2637. doi: 10.1093/nar/gkv133

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© The Author(s) 2015. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 3. — NTP misincorporation induces RNAPII pausing and enhances termination by Rat1/Rai1. (A) Schematic detail for NTP misincorporation via template misalignment. The RNA strand is shown in sky blue and the DNA strand is in dark blue. (B) EC1 scaffold tested. DNA sequences are shown on top of the quantification graph. An asterisk specifies the incorporation site of an incoming NTP (n position). In a control group lacking nuclease, the addition of non-cognate ATP or GTP generated misincorporated RNA bands via template misalignment (blue arrows, ∼34/35 nt), whereas cognate CTP does not. Notably, Rat1/Rai1 more efficiently terminates RNAPII when non-cognate NTP (ATP or GTP) rather than cognate CTP was added. Addition of UTP to EC1 induces strong pausing of RNAPII that does not support further elongation (red arrow), which results in slight inhibition of termination by Rat1/Rai1. (B) EC2 scaffold tested. Different sequences are shown in red. Rat1/Rai1 more effectively terminates RNAPII when a non-cognate NTP (ATP, CTP or UTP), rather than cognate GTP, was added.