Fig. 2: Measuring co-transcriptional cleavage activity by single-molecule nascent RNA sequencing.

a Schematic of the cleavage index (CI) calculation. Full-length readthrough transcripts (blue) have a 5’ end more than 50 nt upstream of the poly(A) site and a 3’ end extending beyond 50 nt downstream. The 5’ cleavage products (red) have a 5’ end before 50 nt upstream of the poly(A) site and a 3’ end within 50 nt upstream or downstream of the poly(A) site. The CI is calculated by dividing the number (m) of 5’ cleavage products by the total number (n + m) of full-length readthrough transcripts and 5’ cleavage products. b Example with PSBP-1 (AT1G06680) in WT. The upper panel shows full-length readthrough transcripts, and the bottom panel shows 5’ cleavage products. The number of reads and the corresponding CI are shown. c Comparison of CI between mutants and their corresponding WT. Scatter plots comparing the CI in mutants (x-axis) to that in WT (y-axis) across protein-coding genes. Each dot represents a single gene. The Pearson correlation coefficient (R²) is shown. The effect size of the difference in CI between mutant and WT for each gene was quantified using Cliff’s Delta (d), calculated from the mean CI of the replicates. The significance of the overall shift in CI distribution between WT and each mutant was determined using a two-sided t-test on the CI values of all genes. Source data are provided as a Source Data file.