The 230926_EJ_Setbp1_AlternativeSplicing GitHub repository includes the following directories and files: bin/ docker/ Dockerfile: The original dockerfile used to build the initial docker image. Future images were pulled from this initial docker image. Rstudio-prefs.json: JSON file with information on RStudio preferences. conda/ environment.yml: Environment files used to build the Conda environment used for alignment with STAR Solo. data/: Contains all processed data generated during this project. The processed data is deposited on Zenodo (refer to the manuscript for details), which includes descriptions of all included files. doc/: Contains any files that were downloaded/accessed for this study. Data_Sheet_2.xlsx: AS gene list identified in human peripheral blood by Liu et al., 2022. G2MGenes.csv: cell cycle genes converted from human to mouse for Seurat pre-processing. Genes were converted on 240623. GOCC_SMALL_NUCLEAR_RIBONUCLEOPROTEIN_COMPLEX.v2023.2.Hs.tsv: List of known spliceosome components. Accessed on 240208 README: README file for this subdirectory. SGenes.csv: ell cycle genes converted from human to mouse for Seurat pre-processing. Genes were converted on 240623. results/ ambientRNA_removal/: includes rho density plots. as_gene_summaries/: includes publication-ready 3-panel figures for all significant SJU genes brain/: includes 33 significant SJU genes in the cerebral cortex kidney/: includes 62 significant SJU genes in the kidney distributions/: includes 5 PNGs of different gene, splice junction, and PSI score distributions in the cerebral cortex final_outputs/: includes all final figures. These can be accessed through the GitHub repository or seen in the paper. kidney_figures/: includes three kidney plots included in the manuscript. kidney_overview_figure.png setbp1.png setbp1_sj_expr_usage.png kidney_outputs/: includes gene and sj violin plots included as supplementary figure in the manuscript. genes_sjs_percell_violin.png marvel_outputs/: includes intermediate marvel outputs, primarily generated from the cerebral cortex data, seurat_outputs/: includes outputs generated during seurat preprocessing for the cerebral cortex and the kidney supplementary_figures/: includes supplementary figures included in the manuscript. tables/: includes tables that are referenced in the manuscript. upset_plots/: includes significant SJU gene UpSet plots in the cerebral cortex and the kidney README: subdirectory README file src/ ambientRNA-removal/ 01_ambientRNA_removal.R: ambient RNA removal at the gene count level of all samples. 01_ambientRNA_removal.sh: driver for 01_ambientRNA_removal.R figures/ figure_2.Rmd: Code to make manuscript figure 2. figure_3-4.Rmd: Code to make manuscript figures 3 and 4. figure_5.Rmd: Code to make manuscript figure 5. functions.R: Functions script for figure scripts written by Emma F. Jones supp_figure_2.Rmd: Code to make manuscript supplementary figure 2. geom_split_violin.R: Code to make split violin plots mean_expression_celltype.R: Code to calculate cell-type-specific mean expression kidney_overview_figure.R: Code to make supplementary figure of kidney dataset overview. kidney_overview_figure.sh: Driver of kidney_overview_figure.R kidney_marvel_figures.R: Code to make MARVEL figures in the kidney data. Kidney_marvel_figures.sh: Driver of kidney_marvel_figures.R marvel/ 01_format_MARVEL_data.Rmd: Code to format cerebral cortex MARVEL data 02_MARVEL_differential_analysis.Rmd: Code for differential MARVEL analysis in the cerebral cortex 03_analyze_de_genes.Rmd: DEA within MARVEL in the cerebral cortex 04_calc_sj_usage.Rmd: Splice junction usage analysis in the cerebral cortex 05_cell_specific_sj_expr.Rmd: Splice junction expression analysis in the cerebral cortex 06_format_kidney_MARVEL_data.R: Code to format kidney MARVEL data 06_format_kidney_MARVEL_data.sh: Driver of 06_format_kidney_MARVEL_data.R 07_kidney_MARVEL_differential_analysis.R: Code for differential MARVEL analysis in the kidney 07_kidney_MARVEL_differential_analysis.sh: Driver of 07_kidney_MARVEL_differential_analysis.R 08_analyze_kidney_de_genes.R: DEA within MARVEL in the kidney 08_analyze_kidney_de_genes.sh: Driver of 08_analyze_kidney_de_genes.R 09_calc_sj_usage_kidney.R: Splice junction usage analysis in the kidney 09_calc_sj_usage_kidney.sh: Driver of 09_calc_sj_usage_kidney.R 10_cell_specific_kidney_sj_expr.R: Splice junction expression analysis in the kidney 10_cell_specific_kidney_sj_expr.sh: Driver of 10_cell_specific_kidney_sj_expr.R PlotSJPosition_modification.R: Modified MARVEL code to plot SJ position functions.R: MARVEL functions script written by Emma F. Jones samtools/: sort_index_bam.sh: Code to sort the bam indices. seurat/ 01_import_filter_data.Rmd: R Markdown for preprocessing cerebral cortex data 02_annotate_cell_types.Rmd: R Markdown for cell typing of the cerebral cortex data 03_kidney_preprocessing.R: Script for full Seurat preprocessing of the kidney data 03_kidney_preprocessing.sh: Driver of 03_kidney_preprocessing.R functions.R: Seurat-specific functions script written by Emma F. Jones generate_cellcycle_lists.R: R script to generate mouse cell cycle lists for Seurat quality control, written by Timothy C. Howton starsolo_conda/ 01_build_STAR_genome.sh: Script for creating the STAR genome needed for alignment 02_run_STARsolo.sh: Script for running STAR Solo on all included samples. README: Subdirectory-specific README with STAR Solo instructions. README: source directory-specific README file functions_soelter.R: R script with functions for any code pertaining to manuscript revisions. Written and modified by Tabea M. Soelter. verifications/: includes multiple .md5 files that include script specific MD5sums used during verification in internal peer review What's Changed Ambient rna removal by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/91 Seurat processing after soupX by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/92 Marvel re-analysis by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/93 Brain DESeq2 Re-analysis by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/94 Seurat pre-processing of the kidney S858R data by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/95 Figure re-plotting for brain by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/96 MARVEL analysis in kidney by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/97 Kidney DESeq2 Analysis by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/98 Kidney plotting by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/99 Fix brain as summaries by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/100 Clean repo by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/102 Update readme by @tsoelter in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/101 New Contributors @tsoelter made their first contribution in https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/pull/91 Full Changelog: https://github.com/lasseignelab/230926_EJ_Setbp1_AlternativeSplicing/compare/1.0.0...2.0.0