Fig. 5. Single-molecule FRET studies of nascent tbpA aptamers obtained by stepwise transcription.

a Schematic representing the nascent RNA construct. The nascent RNA has been produced using stepwise transcription reactions and contains a 3’ sequence allowing to hybridize to a DNA anchor coupled to a biotin. b smFRET histograms of the nascent tbpA aptamer obtained in the absence and presence of 1 mM TPP. The folded (F) and unfolded (U) states are indicated. Histograms obtained without and with TPP were built using 210 molecules and 398 molecules, respectively. c smFRET time trajectories obtained in the absence (Left) and presence (Right) of TPP. The anti-correlated Cy3 donor and Cy5 acceptor emission intensities are shown with the resulting FRET trace. Photobleaching events are shown by an asterisk. d smFRET contour plots obtained in the absence (Left) and presence (Right) of TPP. Selected traces have been accumulated for the first 50 s. Histograms representing the percentual occupancy of each state are shown to the right. Contour plots obtained without and with TPP were built using 108 molecules and 117 molecules, respectively.