This repository contains the code used for generating the figures depicted in the following paper: Quantitative analysis of C. elegans transcripts by Nanopore direct cDNA sequencing reveals terminal hairpins in non trans- spliced mRNAs. Bernard, et al. 2022.
The manuscript is available on BioRxiv.
- Florian Bernard ¹𝄒²
- Delphine Dargere ¹
- Oded Rechavi ²
- Denis Dupuy ¹
¹ Université de Bordeaux, Inserm U1212, CNRS UMR5320 , Institut Européen de Chimie et Biologie (IECB), 2, rue Robert Escarpit, 33607 Pessac, France.
² Department of Neurobiology, Wise Faculty of Life Sciences & Sagol School of Neuroscience, Tel Aviv University, Tel Aviv, Israel.
The direct-cDNA datasets used in this study have been deposited in the Sequence Read Archive (SRA) under the following accession code: PRJNA822363.
The processed dataset, along with the necessary files for running the notebooks locally have been deposited on Figshare (DOI: 10.6084/m9.figshare.19131260).
A preprocessing notebook is available for generating the dataset table used in all downstream analysis from SAM/BAM alignments files (retrieved from our SRA archive or for analyzing your own alignment files).
A separate notebook was then generated for each of the figures shown in the paper as detailed above:
| Notebook | Figure | Title |
|---|---|---|
| pre-processing | - | Pipeline for processing alignments and generating the dataset table |
| Fig1_notebook | Fig. 1b | Measure of base quality in 5' soft-clip region and alignment region |
| Fig2_notebook | Fig. 2b | Measure of total SL1 and SL2 variants detected in our reads |
| Fig3_notebook | Fig. 3a | Spliced leader usage |
| Fig3_notebook | Fig. 3b | SL2 gene specificity |
| Fig4_notebook | Fig. 4a | Gene expression and trans-splicing status |
| Fig4_notebook | Fig. 4b | Trans-splicing detection level |
| Fig4_notebook | Fig. 4c | Poorly trans spliced mRNA have a propensity to form a 5’ stem loop structure |
| Fig5_notebook | Fig. 5a | Measure of ratio SL/Hairpin |
| Fig5_notebook | Fig. 5b | Proportion of trans-spliced gene with various SL thresholds |
| Fig5_notebook | Fig. 5c | Read coverage for SL, Hairpin and Unidentified genes |
| Fig6_notebook | Fig. 6 | Schematic representation of the trans-splicing information |
| SupFig1_notebook | Sup. Fig. 1 | Measure of strand bias in direct-cDNA experiments |
| SupFig2_notebook | Sup. Fig. 2b | Measured length of 5’ and 3’ soft-clips |
| SupFig3_notebook | Sup. Fig. 3a | Quantification of sequencing adapters in soft-clips regions |
| SupFig3_notebook | Sup. Fig. 3b | Origin of supplementary alignments |
| SupFig3_notebook | Sup. Fig. 3c | Size distribution of supplementary alignments |
| SupFig4_notebook | Sup. Fig. 4 | Measure of base quality across all sequencing experiments |
| SupFig5_notebook | Sup. Fig. 5a | Measure of read length and alignment length across experiments |
| SupFig5_notebook | Sup. Fig. 5b | Measure of read coverage |
| SupFig6_notebook | Sup. Fig. 6a | Strand orientation for unidentified reads |
| SupFig6_notebook | Sup. Fig. 6b | Length of 5’ soft-clips versus their alignment length |
| SupFig7_notebook | Sup. Fig. 7a | Single SL1 promotor - nlp-36 (B0464.3) |
| SupFig7_notebook | Sup. Fig. 7b | Multiple SL1 promotors - M60.4 |
| SupFig7_notebook | Sup. Fig. 7c | Operon organization - rla-1 (Y37E3.8) & Y37E3.7 |
| SupFig7_notebook | Sup. Fig. 7d | Differentially trans-spliced promoters - lev-11 (Y105E8B.1) |
| SupFig8_notebook | Sup. Fig. 8a | Measure of strand bias at each locus in SSP experiments |
| SupFig8_notebook | Sup. Fig. 8b | Unbiased locus presents a high concentration of SSP reads |
| SupFig8_notebook | Sup. Fig. 8c | Unbiased locus in SSP Exp. are found biased in SL1/NP Exp |
| SupFig9_notebook | Sup. Fig. 9 | Method for evaluating base quality |
| SupFig10_notebook | Sup. Fig. 10 | high confidence SL matchs are located near the alignment start |
This web-app allows you to generate and save plots for your genes of interests without having to run the notebooks.
The app was made using the streamlit library (see streamlit_app.py) and is hosted via streamlit sharing.
The source code is licensed under the MIT License.
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