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. 2021 Jun 16;49(12):7122–7138. doi: 10.1093/nar/gkab462

Figure 7.

Figure 7.

smFRET measurements of pseudoknot stability. (A) Labeling strategy for smFRET experiments. DONGV DB RNA was site-specifically labeled with Cy3. A 3′ extension on the RNA was used to immobilize the RNA construct on a microscope slide based on hybridization to a biotinylated and Cy5-labeled DNA handle. A folded pseudoknot is predicted to have a high FRET value and an unfolded pseudoknot is predicted to have a lower FRET value. (B) smFRET histogram of wild-type DB RNA. (left) Model of the RNA construct. (right) Histogram of the distribution of FRET values, n = number of molecules observed. (C) Representative smFRET trace of a wild-type DB RNA. (D) smFRET histogram of PKmut DB RNA. (left) Model of the RNA construct used. (right) smFRET histogram. (E) Representative smFRET trace of a PKmut DB RNA.