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--- a +++ b/settings.R @@ -0,0 +1,317 @@ +suppressPackageStartupMessages(library(SingleCellExperiment)) +suppressPackageStartupMessages(library(data.table)) +suppressPackageStartupMessages(library(purrr)) +suppressPackageStartupMessages(library(ggplot2)) +suppressPackageStartupMessages(library(ggpubr)) +suppressPackageStartupMessages(library(stringr)) +suppressPackageStartupMessages(library(argparse)) + +######### +## I/O ## +######### + +io <- list() +if (grepl("ricard",Sys.info()['nodename'])) { + io$basedir <- "/Users/ricard/data/gastrulation_multiome_10x" + io$atlas.basedir <- "/Users/ricard/data/gastrulation10x" + io$gene_metadata <- "/Users/ricard/data/ensembl/mouse/v87/BioMart/all_genes/Mmusculus_genes_BioMart.87.txt" +} else if (Sys.info()[['nodename']]=="BI2404M") { + io$basedir <- "/Users/argelagr/data/gastrulation_multiome_10x/test" + io$atlas.basedir <- "/Users/argelagr/data/gastrulation10x" + io$gene_metadata <- "/Users/argelagr/data/ensembl/mouse/v87/BioMart/mRNA/Mmusculus_genes_BioMart.87.txt" + io$TFs_file <- "/Users/argelagr/data/mm10_regulation/TFs/TFs.txt" +} else if (grepl("pebble|headstone", Sys.info()['nodename'])) { + if (grepl("Clark", Sys.info()['effective_user'])) { + io$basedir <- "/bi/scratch/Stephen_Clark/multiome/resilio" + io$rawdata <- "/bi/scratch/Stephen_Clark/multiome/raw" + io$gene_metadata <- "/bi/scratch/Stephen_Clark/annotations/Mmusculus_genes_BioMart.87.txt" + } else if (grepl("argelag", Sys.info()['effective_user'])) { + io$basedir <-'/bi/group/reik/ricard/data/gastrulation_multiome_10x/test' + io$archR.directory <- file.path(io$basedir,"processed/atac/archR") + io$gene_metadata <- "/bi/group/reik/ricard/data/ensembl/mouse/v87/BioMart/all_genes/Mmusculus_genes_BioMart.87.txt" + io$atlas.basedir <- "/bi/group/reik/ricard/data/pijuansala2019_gastrulation10x" + io$TFs_file <- "/bi/group/reik/ricard/data/mm10_regulation/TFs/TFs.txt" + } +} else if (grepl("bi2228m", Sys.info()['nodename'])) { + io$basedir <- "/Users/clarks/data/multiome/" + io$rawdata <- "/Users/clarks/data/multiome_raw_sub/" +} else if(grepl('LAPTOP',Sys.info()['nodename'])){ + io$basedir <- 'D:/OneDrive - zju.edu.cn/research/bioin/lab/Ricard/data/gastrulation_multiome_10x' +} else if(grepl('Workstation',Sys.info()['nodename'])){ + io$basedir <-'/home/lijingyu/gastrulation/data/gastrulation_multiome_10x' + io$gene_metadata <- "/home/lijingyu/gastrulation/data/gastrulation_multiome_10x/Mmusculus_genes_BioMart.87.txt" + io$atlas.basedir <- "/home/lijingyu/gastrulation/data/gastrulation10x" +} else { + stop("Computer not recognised") +} + +io$metadata <- file.path(io$basedir,"sample_metadata.txt.gz") + +# TFs +# io$TFs <- file.path(io$basedir,"results/TFs.txt") + +# RNA +io$rna.anndata <- file.path(io$basedir,"processed/rna/anndata.h5ad") +io$rna.seurat <- file.path(io$basedir,"processed/rna/seurat.rds") +io$rna.sce <- file.path(io$basedir,"processed/rna/SingleCellExperiment.rds") +io$rna.tfs.sce <- file.path(io$basedir,"processed/rna/SingleCellExperiment_TFs.rds") +io$rna.pseudobulk.sce <- file.path(io$basedir,"results/rna/pseudobulk/celltype/SingleCellExperiment_pseudobulk.rds") +io$rna.tfs.pseudobulk.sce <- file.path(io$basedir,"results/rna/pseudobulk/celltype/SingleCellExperiment_TFs_pseudobulk.rds") +io$rna.metacells.sce <- file.path(io$basedir,"results/rna/metacells/all_cells/SingleCellExperiment_metacells.rds") + +# Differential RNA expression and marker genes +# io$rna.differential <- file.path(io$basedir,"results/rna/differential") +io$rna.celltype.marker_genes.all <- file.path(io$basedir,"results/rna/differential/pseudobulk/celltype/parsed/marker_genes_all.txt.gz") +io$rna.celltype.marker_genes.filt <- file.path(io$basedir,"results/rna/differential/pseudobulk/celltype/parsed/marker_genes_filtered.txt.gz") +io$rna.celltype.marker_tfs.all <- file.path(io$basedir,"results/rna/differential/pseudobulk/celltype/parsed/marker_tfs_all.txt.gz") +io$rna.celltype.marker_tfs.filt <- file.path(io$basedir,"results/rna/differential/pseudobulk/celltype/parsed/marker_tfs_filtered.txt.gz") + +# RNA atlas (Pijuan-Sala2019) +io$rna.atlas.metadata <- file.path(io$atlas.basedir,"sample_metadata.txt.gz") +io$rna.atlas.sce <- file.path(io$atlas.basedir,"processed/SingleCellExperiment.rds") +io$rna.atlas.marker_genes.up <- file.path(io$atlas.basedir,"results/differential/celltypes/genes/all_stages/marker_genes/marker_genes_upregulated_filtered.txt.gz") +io$rna.atlas.marker_genes.all <- file.path(io$atlas.basedir,"results/differential/celltypes/genes/all_stages/marker_genes/marker_genes_upregulated_all.txt.gz") +io$rna.atlas.marker_TFs.up <- file.path(io$atlas.basedir,"results/differential/celltypes/TFs/TF_markers/marker_TFs_upregulated_filtered.txt.gz") +io$rna.atlas.marker_TFs.all <- file.path(io$atlas.basedir,"results/differential/celltypes/TFs/TF_markers/marker_TFs_upregulated_all.txt.gz") +io$rna.atlas.differential <- file.path(io$atlas.basedir,"results/differential") +# io$rna.atlas.average_expression_per_celltype <- file.path(io$atlas.basedir,"results/marker_genes/all_stages/avg_expr_per_celltype_and_gene.txt.gz") +io$rna.atlas.sce.pseudobulk <- file.path(io$atlas.basedir,"results/pseudobulk/SingleCellExperiment_pseudobulk.rds") +io$rna.atlas.celltype_proportions <- file.path(io$atlas.basedir,"results/celltype_proportions/celltype_proportions.txt.gz") + +# ATAC +# io$signac.directory <- file.path(io$basedir,"processed/atac/signac") +# io$signac <- file.path(io$signac.directory,"/signac.rds") +# io$cistopic.model <- file.path(io$basedir,"results/atac/cistopic/cistopic_warpLDA_bestModel.rds") +io$archR.directory <- file.path(io$basedir,"processed/atac/archR") +io$motifmatcher_scores.se <- file.path(io$archR.directory,"Annotations/CISBP-Scores.rds") +io$motifmatcher_positions.se <-file.path(io$archR.directory,"Annotations/CISBP-Positions.rds") +io$archR.projectMetadata <- file.path(io$archR.directory,"projectMetadata.rds") +io$archR.peakSet.granges <- file.path(io$archR.directory,"PeakSet.rds") +io$archR.peakSet.stats <- file.path(io$basedir,"results/atac/archR/feature_stats/PeakMatrix/PeakMatrix_celltype_stats.txt.gz") +io$archR.bgdPeaks <- file.path(io$archR.directory,"Background-Peaks.rds") +io$archR.peakSet.bed <- file.path(io$archR.directory,"PeakCalls/peaks_archR_macs2.bed.gz") +io$archR.peakMatrix.cells <- file.path(io$archR.directory,"Matrices/PeakMatrix_summarized_experiment.rds") +io$archR.peakMatrix.metacells <- file.path(io$basedir,"results/atac/archR/metacells/all_cells/PeakMatrix/PeakMatrix_summarized_experiment_metacells.rds") +io$archR.peakMatrix.pseudobulk <- file.path(io$basedir,"results/atac/archR/pseudobulk/celltype/PeakMatrix/pseudobulk_PeakMatrix_summarized_experiment.rds") +io$archR.peak.metadata <- file.path(io$archR.directory,"PeakCalls/peak_metadata.tsv.gz") +io$archR.peak.stats <- file.path(io$basedir,"results/atac/archR/feature_stats/PeakMatrix_celltype.mapped_stats.txt.gz") +io$archR.peak2gene.all <- file.path(io$basedir,"results/atac/archR/peak_calling/peaks2genes/peaks2genes_all.txt.gz") +io$archR.peak2gene.nearest <- file.path(io$basedir,"results/atac/archR/peak_calling/peaks2genes/peaks2genes_nearest.txt.gz") +io$archR.GeneScoreMatrix.cells <- file.path(io$basedir,"processed/atac/archR/Matrices/GeneScoreMatrix_TSS_summarized_experiment.rds") +io$archR.GeneScoreMatrix.pseudobulk <- file.path(io$basedir,"results/atac/archR/pseudobulk/celltype/GeneScoreMatrix_TSS/pseudobulk_GeneScoreMatrix_TSS_summarized_experiment.rds") +io$archR.chromvar.cells <- file.path(io$basedir,"results/atac/archR/chromvar/cells/chromVAR_deviations_CISBP_archr.rds") +io$archR.chromvar.pseudobulk <- file.path(io$basedir,"results/atac/archR/chromvar/pseudobulk/celltype/chromVAR_deviations_CISBP_pseudobulk_archr.rds") +io$archR.feature_stats <- file.path(io$basedir,"results/atac/archR/chromvar_chip/pseudobulk/chromVAR_chip_CISBP_archr.rds") +io$archR.motif2gene <- file.path(io$basedir,"processed/atac/archR/Annotations/CISBP_motif2gene.txt.gz") +# io$archR.chromvar_chip.cells <- file.path(io$basedir,"results/atac/archR/chromvar/cells/celltype/chromVAR_deviations_CISBP_pseudobulk_archr.rds") + +# io$archR.pseudobulk.genes.se <- file.path(io$basedir,"processed/atac/archR/pseudobulk/celltype.predicted/pseudobulk_GeneScoreMatrix_summarized_experiment.rds") + +# Differential ATAC and marker peaks +io$atac.markers_peaks.pseudobulk.all <- file.path(io$basedir,"results/atac/archR/differential/pseudobulk/celltype/PeakMatrix/parsed/markers_all.txt.gz") +io$atac.markers_peaks.pseudobulk.filt <- file.path(io$basedir,"results/atac/archR/differential/pseudobulk/celltype/PeakMatrix/parsed/markers_filt.txt.gz") +# io$archR.markers_chromvar <- file.path(io$basedir,"results/atac/archR/chromvar/differential/markers/marker_TFs.txt.gz") + +# ATAC public data (Pijuan-Sala2020) +io$pijuansala.basedir <- file.path(io$basedir,"public_datasets/Pijuan-Sala_2020") +io$pijuansala.archR.directory <- file.path(io$pijuansala.basedir,"data/processed/archR") + +# paga +io$paga.connectivity <- file.path(io$atlas.basedir,"results/paga/paga_connectivity.csv") +io$paga.coordinates <- file.path(io$atlas.basedir,"results/paga/paga_coordinates.csv") + +# Virtual ChIP-seq +# io$tf2peak_cor.se <- file.path(io$basedir,"results/rna_atac/rna_vs_acc/pseudobulk/TFexpr_vs_peakAcc/cor_TFexpr_vs_peakAcc_SummarizedExperiment.rds") +# io$tf.activities <- file.path(io$atlas.basedir,"results/rna_atac/rna_vs_chromvar/TF_activities/tf_activities.txt.gz") +io$virtual_chip.dir <- file.path(io$basedir,"results/rna_atac/virtual_chipseq") +io$virtual_chip.mtx <- file.path(io$basedir,"results/rna_atac/virtual_chipseq/virtual_chip_mtx.rds") +io$archR.chromvar_chip.pseudobulk <- file.path(io$basedir,"results/atac/archR/chromvar_chip/pseudobulk/chromVAR_chip_CISBP_archr.rds") + +# Coexpression +# io$tf2tf_cor.mtx <- file.path(io$basedir,"results/rna/coexpression/correlation_matrix_tf2tf.rds") +# io$tf2gene_cor.mtx <- file.path(io$basedir,"results/rna/coexpression/correlation_matrix_tf2gene.rds") + +# Dimensionality reduction +io$rna.dimred.pca <- file.path(io$basedir,"results/rna/dimensionality_reduction/sce/batch_correction_sample_remove_ExE_cells_False/pca_features2500_pcs50.txt.gz") +io$atac.dimred.lsi <- file.path(io$basedir,"results/atac/archR/dimensionality_reduction/cells/PeakMatrix/remove_ExE_cells_False/batch_correction_None/lsi_nfeatures25000_ndims50.txt.gz") +# io$rna.dimred.umap <- file.path(io$basedir,"results/rna/dimensionality_reduction/all_cells/E7.5_rep1-E7.5_rep2-E8.0_rep1-E8.0_rep2-E8.5_rep1-E8.5_rep2_umap_features2500_pcs30_neigh25_dist0.3.txt.gz") +# io$atac.dimred.umap <- file.path(io$basedir,"results/atac/archR/dimensionality_reduction/PeakMatrix/all_cells/E7.5_rep1-E7.5_rep2-E8.0_rep1-E8.0_rep2-E8.5_rep1-E8.5_rep2_umap_nfeatures50000_ndims50_neigh30_dist0.45.txt.gz") + +############# +## Options ## +############# + +opts <- list() + +opts$stages <- c( + "E7.5", + "E7.75", + "E8.0", + "E8.5", + "E8.75" +) + +opts$samples <- c( + "E7.5_rep1", + "E7.5_rep2", + "E7.75_rep1", + "E8.0_rep1", + "E8.0_rep2", + "E8.5_rep1", + "E8.5_rep2", + "E8.5_CRISPR_T_WT", + "E8.5_CRISPR_T_KO", + "E8.75_rep1", + "E8.75_rep2" +) + +# opts$rename.samples <- c( +# "E8.75_rep1" = "e8_75_1_L002", +# "E8.75_rep2" = "e8_75_2_L002", +# "E8.5_rep1" = "multiome1", +# "E8.5_rep2" = "multiome2", +# "E8.0_rep1" = "E8_0_rep1_multiome", +# "E8.0_rep2" = "E8_0_rep2_multiome", +# "E7.5_rep1" = "rep1_L001_multiome", +# "E7.5_rep2" = "rep2_L002_multiome" +# # E7.75_rep1 +# # E8.5_CRISPR_T_WT +# # E8.5_CRISPR_T_KO +# ) + +opts$sample2stage <- c( + "E7.5_rep1" = "E7.5", + "E7.5_rep2" = "E7.5", + "E7.75_rep1" = "E7.75", + "E8.0_rep1" = "E8.0", + "E8.0_rep2" = "E8.0", + "E8.5_rep1" = "E8.5", + "E8.5_rep2" = "E8.5", + "E8.75_rep1" = "E8.75", + "E8.75_rep2" = "E8.75", + "E8.5_CRISPR_T_KO" = "E8.5", + "E8.5_CRISPR_T_WT" = "E8.5" +) + + +opts$celltypes <- c( + "Epiblast", + "Primitive_Streak", + "Caudal_epiblast", + "PGC", + "Anterior_Primitive_Streak", + "Notochord", + "Def._endoderm", + "Gut", + "Nascent_mesoderm", + "Mixed_mesoderm", + "Intermediate_mesoderm", + "Caudal_Mesoderm", + "Paraxial_mesoderm", + "Somitic_mesoderm", + "Pharyngeal_mesoderm", + "Cardiomyocytes", + "Allantois", + "ExE_mesoderm", + "Mesenchyme", + "Haematoendothelial_progenitors", + "Endothelium", + "Blood_progenitors_1", + "Blood_progenitors_2", + "Erythroid1", + "Erythroid2", + "Erythroid3", + "NMP", + "Rostral_neurectoderm", + "Caudal_neurectoderm", + "Neural_crest", + "Forebrain_Midbrain_Hindbrain", + "Spinal_cord", + "Surface_ectoderm", + "Visceral_endoderm", + "ExE_endoderm", + "ExE_ectoderm", + "Parietal_endoderm" +) + +opts$stage.colors = c( + "E7.5" = "#edf8b1", + "E7.75" = "#c7e9b4", + "E8.0" = "#7fcdbb", + "E8.25" = "#41b6c4", + "E8.5" = "#1d91c0", + "E8.75" = "#225ea8" +) +# opts$stage.colors <- viridis::viridis(n=length(opts$stages)) +# names(opts$stage.colors) <- rev(opts$stages) + +opts$celltype.colors = c( + "Epiblast" = "#635547", + "Primitive_Streak" = "#DABE99", + "Caudal_epiblast" = "#9e6762", + "PGC" = "#FACB12", + "Anterior_Primitive_Streak" = "#c19f70", + "Notochord" = "#0F4A9C", + "Def._endoderm" = "#F397C0", + "Gut" = "#EF5A9D", + "Nascent_mesoderm" = "#C594BF", + "Mixed_mesoderm" = "#DFCDE4", + "Intermediate_mesoderm" = "#139992", + "Caudal_Mesoderm" = "#3F84AA", + "Paraxial_mesoderm" = "#8DB5CE", + "Somitic_mesoderm" = "#005579", + "Pharyngeal_mesoderm" = "#C9EBFB", + "Cardiomyocytes" = "#B51D8D", + "Allantois" = "#532C8A", + "ExE_mesoderm" = "#8870ad", + "Mesenchyme" = "#cc7818", + "Haematoendothelial_progenitors" = "#FBBE92", + "Endothelium" = "#ff891c", + "Blood_progenitors" = "#c9a997", + "Blood_progenitors_1" = "#f9decf", + "Blood_progenitors_2" = "#c9a997", + "Erythroid" = "#EF4E22", + "Erythroid1" = "#C72228", + "Erythroid2" = "#f79083", + "Erythroid3" = "#EF4E22", + "NMP" = "#8EC792", + "Neurectoderm" = "#65A83E", + "Rostral_neurectoderm" = "#65A83E", + "Caudal_neurectoderm" = "#354E23", + "Neural_crest" = "#C3C388", + "Forebrain_Midbrain_Hindbrain" = "#647a4f", + "Spinal_cord" = "#CDE088", + "Surface_ectoderm" = "#f7f79e", + "Visceral_endoderm" = "#F6BFCB", + "ExE_endoderm" = "#7F6874", + "ExE_ectoderm" = "#989898", + "Parietal_endoderm" = "#1A1A1A" +) + +opts$chr <- paste0("chr",c(1:19,"X","Y")) + +################### +## Load metadata ## +################### + +# sample_metadata <- fread(io$metadata) +# .[pass_QC==T] %>% +# .[batch%in%opts$batches] %>% +# .[,celltype.mapped:=stringr::str_replace_all(celltype.mapped," ","_")] %>% +# .[,celltype.mapped:=stringr::str_replace_all(celltype.mapped,"/","_")] %>% +# .[,celltype.mapped:=factor(celltype.mapped, levels=names(opts$celltype.colors))] + + +################### +## Edit metadata ## +################### + +# io$metadata <- file.path(io$basedir,"results/atac/archR/qc/sample_metadata_after_qc.txt.gz") +# io$metadata <- file.path(io$basedir,"results/rna/mapping/sample_metadata_after_mapping.txt.gz") +# sample_metadata <- fread(io$metadata) +# sample_metadata[pass_rnaQC==FALSE & nFrags_atac<=1e4,pass_atacQC:=FALSE] +# sample_metadata[sample=="E7.75_rep1" & ribosomal_percent_RNA>=8,pass_rnaQC:=FALSE] +# sample_metadata[sample=="E7.75_rep1" & ribosomal_percent_RNA>=4.5 & nFeature_RNA<=3500,pass_rnaQC:=FALSE] +# fwrite(sample_metadata, io$metadata, sep="\t", quote=F, na="NA") + +