Fig. 2: Promoted read-through efficiencies and mutations against CMC-Ψ by RT-1306.

a Shown on the left are the fluorescence images of RT stop assay gels for SSIII, wtRT, and RT-1306 reading against Ψ-oligo1 and Ψ-oligo2 RNAs with and without CMC treatment. RT products were run and separated on 15% 8 M Urea-PAGE gels and imaged by the fluorescence imaging. Positions of the FAM-labeled RT primer, truncated cDNA at the Ψ site, and the full-length cDNA products are labeled with “P”, “T”, and “FL”, respectively. The same gels were also stained by SYBR-Gold and imaged (Supplementary Fig. 2). Shown on the right are quantified RT read-through based on the RT stop assay. The RT read-through efficiency over CMC-Ψ were quantified by the ratio of the fluorescence intensity of the “T” band over the sum of intensities of the “T” and “FL” bands. Error bars represent the standard deviations of n = 3 replicates for Ψ-oligo1, n = 2 replicates for Ψ-oligo2; full gel images of 2 replicates are presented in Supplementary Fig. 2. b Colony sequencing data of the cDNA products from RT-1306 processing Ψ-oligo1 and Ψ-oligo2 RNAs with (“CMC+”) and without (“CMC−”) CMC treatments (Methods). Shown are the clones that carry mutations at the Ψ sites and data for all sequenced colonies are shown in Supplementary Fig. 3.