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. 2021 Dec 6;49(22):13179–13193. doi: 10.1093/nar/gkab1170

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Published by Oxford University Press on behalf of Nucleic Acids Research 2021.

This work is written by (a) US Government employee(s) and is in the public domain in the US.

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Figure 3. — Formation and stability of modified ^GCPAN triplex. (A) Secondary structures of the unimolecular ^GCPAN triplex (left) and bimolecular ^GCENE•ggc(A)₉ triplex (right). (B) Susceptibility of the unimolecular ^GCPAN triplex to 3′–5′ exoribonuclease R (RNaseR) digestion evaluated by denaturing PAGE. (C) CD spectra for ggc(A)₉ (black), ^GCENE (red) and ^GCENE+ggc(A)₉ mixture (blue). Distinct spectra for each sample indicate triple association in the ^GCENE + ggc(A)₉ mixture. (D) SEC analysis indicates enhanced stability of the ^GCPAN triplex. The main SEC fraction was analyzed by denaturing polyacrylamide gel electrophoresis, indicating the presence of ggc(A)₉ and ^GCENE duplex. Bands corresponding to ^GCENE duplex in the control sample (left lane) and main SEC fraction (right lane) are oversaturated, resulting in a shadow effect and an appearance as a light orange band. (E) MST binding plot for ^GCENE titrated into 5′-Cy5 labeled ggc(A)₉ indicates high affinity binding of the two RNA strands. (F) EMSA analysis of binding of ^GCENE duplex titrated into ³²P-radiolabeled ggc(A)₉ also indicates high binding affinity for these two RNA strands.