Figure - PMC

Skip to main content

An official website of the United States government

Here's how you know

Here's how you know

Official websites use .gov
A .gov website belongs to an official government organization in the United States.

Secure .gov websites use HTTPS
A lock ( ) or https:// means you've safely connected to the .gov website. Share sensitive information only on official, secure websites.

View full-text article in PMC

. 2024 Sep 11;26(10):1700–1711. doi: 10.1038/s41556-024-01493-w

Search in PMC
Search in PubMed
View in NLM Catalog
Add to search

© The Author(s) 2024

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.

PMC Copyright notice

Extended Data Fig. 3 — (a) MCP-GFP expression is uniform across the cell population correlated with DNA content (DAPI signal) (n = 1). (b) (Left panel) Measurements of GFP photobleaching (grey datapoints) over a full time-course of live-cell-imaging approximated with an exponential decay (red line) that was used to correct fluorescence intensity in time-course transcription trajectories. (Right panel) Examples of the effect of this correction are presented on the right. (c) To measure the intensity of single pre-mRNAs containing 128 MS2 aptamers, imaging was performed using a higher 490nm excitation intensity. The curve quantifies MCP-GFP intensity (y-axis) in response to varying 490nm excitation levels. The blue dashed lines represent values used for live-cell transcription imaging and for single pre-mRNA intensity quantification (dashed line with arrow-head). This curve informed us of the 490nm intensity that excites GFP at 3x the value used in our live-cell transcription measurements. (d) Histograms of single pre-mRNA intensities recalculated in values corresponding to live-cell transcription measurements for Zic2, E2f6, and Hspg2. The red line represents a Gaussian fit with mean and standard deviation values indicated above. These values allowed us to recalculate fluorescence intensity units in order to attribute transcript numbers based on fluorescence intensity at the transcription site. Data represents single biological replicate. (e) Examples of live-cell transcription trajectories with identified ON-periods indicated in blue or orange depending on whether they were taken into account during RNA Pol II reinitiation rate estimations or not. All ON-periods were taken into account in amplitude and duration analysis. (f) An example of a live-cell transcription trajectory with three ON-periods (in blue) with their amplitudes and RNA PolII reinitiation rates (from linear fits, red dashed lines) indicated. Source numerical data are available in source data.

Source data