Transgene expression
Katharina Hembach
5/17/2021
Last updated: 2021-05-19
Checks: 7 0
Knit directory: neural_scRNAseq/
This reproducible R Markdown analysis was created with workflowr (version 1.6.2). The Checks tab describes the reproducibility checks that were applied when the results were created. The Past versions tab lists the development history.
Great! Since the R Markdown file has been committed to the Git repository, you know the exact version of the code that produced these results.
Great job! The global environment was empty. Objects defined in the global environment can affect the analysis in your R Markdown file in unknown ways. For reproduciblity it's best to always run the code in an empty environment.
The command set.seed(20200522) was run prior to running the code in the R Markdown file. Setting a seed ensures that any results that rely on randomness, e.g. subsampling or permutations, are reproducible.
Great job! Recording the operating system, R version, and package versions is critical for reproducibility.
Nice! There were no cached chunks for this analysis, so you can be confident that you successfully produced the results during this run.
Great job! Using relative paths to the files within your workflowr project makes it easier to run your code on other machines.
Great! You are using Git for version control. Tracking code development and connecting the code version to the results is critical for reproducibility.
The results in this page were generated with repository version e616506. See the Past versions tab to see a history of the changes made to the R Markdown and HTML files.
Note that you need to be careful to ensure that all relevant files for the analysis have been committed to Git prior to generating the results (you can use wflow_publish or wflow_git_commit). workflowr only checks the R Markdown file, but you know if there are other scripts or data files that it depends on. Below is the status of the Git repository when the results were generated:
Ignored files:
Ignored: .DS_Store
Ignored: .Rhistory
Ignored: .Rproj.user/
Ignored: ._.DS_Store
Ignored: ._Filtered.pdf
Ignored: ._Rplots.pdf
Ignored: ._Unfiltered.pdf
Ignored: .__workflowr.yml
Ignored: ._coverage.pdf
Ignored: ._coverage_sashimi.pdf
Ignored: ._coverage_sashimi.png
Ignored: ._neural_scRNAseq.Rproj
Ignored: ._pbDS_cell_level.pdf
Ignored: ._pbDS_top_expr_umap.pdf
Ignored: ._pbDS_upset.pdf
Ignored: ._sashimi.pdf
Ignored: ._stmn2.pdf
Ignored: ._tdp.pdf
Ignored: analysis/.DS_Store
Ignored: analysis/.Rhistory
Ignored: analysis/._.DS_Store
Ignored: analysis/._01-preprocessing.Rmd
Ignored: analysis/._01-preprocessing.html
Ignored: analysis/._02.1-SampleQC.Rmd
Ignored: analysis/._03-filtering.Rmd
Ignored: analysis/._04-clustering.Rmd
Ignored: analysis/._04-clustering.knit.md
Ignored: analysis/._04.1-cell_cycle.Rmd
Ignored: analysis/._05-annotation.Rmd
Ignored: analysis/._07-cluster-analysis-all-timepoints.Rmd
Ignored: analysis/._Lam-0-NSC_no_integration.Rmd
Ignored: analysis/._Lam-01-NSC_integration.Rmd
Ignored: analysis/._Lam-02-NSC_annotation.Rmd
Ignored: analysis/._NSC-1-clustering.Rmd
Ignored: analysis/._NSC-2-annotation.Rmd
Ignored: analysis/.__site.yml
Ignored: analysis/._additional_filtering.Rmd
Ignored: analysis/._additional_filtering_clustering.Rmd
Ignored: analysis/._index.Rmd
Ignored: analysis/._organoid-01-1-qualtiy-control.Rmd
Ignored: analysis/._organoid-01-clustering.Rmd
Ignored: analysis/._organoid-02-integration.Rmd
Ignored: analysis/._organoid-03-cluster_analysis.Rmd
Ignored: analysis/._organoid-04-group_integration.Rmd
Ignored: analysis/._organoid-04-stage_integration.Rmd
Ignored: analysis/._organoid-05-group_integration_cluster_analysis.Rmd
Ignored: analysis/._organoid-05-stage_integration_cluster_analysis.Rmd
Ignored: analysis/._organoid-06-1-prepare-sce.Rmd
Ignored: analysis/._organoid-06-conos-analysis-Seurat.Rmd
Ignored: analysis/._organoid-06-conos-analysis-function.Rmd
Ignored: analysis/._organoid-06-conos-analysis.Rmd
Ignored: analysis/._organoid-06-group-integration-conos-analysis.Rmd
Ignored: analysis/._organoid-07-conos-visualization.Rmd
Ignored: analysis/._organoid-07-group-integration-conos-visualization.Rmd
Ignored: analysis/._organoid-08-conos-comparison.Rmd
Ignored: analysis/._organoid-0x-sample_integration.Rmd
Ignored: analysis/01-preprocessing_cache/
Ignored: analysis/02-1-SampleQC_cache/
Ignored: analysis/02-quality_control_cache/
Ignored: analysis/02.1-SampleQC_cache/
Ignored: analysis/03-filtering_cache/
Ignored: analysis/04-clustering_cache/
Ignored: analysis/04.1-cell_cycle_cache/
Ignored: analysis/05-annotation_cache/
Ignored: analysis/06-clustering-all-timepoints_cache/
Ignored: analysis/07-cluster-analysis-all-timepoints_cache/
Ignored: analysis/CH-test-01-preprocessing_cache/
Ignored: analysis/Lam-01-NSC_integration_cache/
Ignored: analysis/Lam-02-NSC_annotation_cache/
Ignored: analysis/NSC-1-clustering_cache/
Ignored: analysis/NSC-2-annotation_cache/
Ignored: analysis/TDP-01-preprocessing_cache/
Ignored: analysis/TDP-02-quality_control_cache/
Ignored: analysis/TDP-03-filtering_cache/
Ignored: analysis/TDP-04-clustering_cache/
Ignored: analysis/TDP-05-00-filtering-plasmid-QC_cache/
Ignored: analysis/TDP-05-plasmid_expression_cache/
Ignored: analysis/TDP-06-cluster_analysis_cache/
Ignored: analysis/TDP-07-01-STMN2_expression_cache/
Ignored: analysis/TDP-07-cluster_12_cache/
Ignored: analysis/TDP-08-00-clustering-HA-D96_cache/
Ignored: analysis/TDP-08-01-HA-D96-expression-changes_cache/
Ignored: analysis/TDP-08-02-TDP_target_genes_cache/
Ignored: analysis/TDP-08-clustering-timeline-HA_cache/
Ignored: analysis/additional_filtering_cache/
Ignored: analysis/additional_filtering_clustering_cache/
Ignored: analysis/organoid-01-1-qualtiy-control_cache/
Ignored: analysis/organoid-01-clustering_cache/
Ignored: analysis/organoid-02-integration_cache/
Ignored: analysis/organoid-03-cluster_analysis_cache/
Ignored: analysis/organoid-04-group_integration_cache/
Ignored: analysis/organoid-04-stage_integration_cache/
Ignored: analysis/organoid-05-group_integration_cluster_analysis_cache/
Ignored: analysis/organoid-05-stage_integration_cluster_analysis_cache/
Ignored: analysis/organoid-06-conos-analysis_cache/
Ignored: analysis/organoid-06-conos-analysis_test_cache/
Ignored: analysis/organoid-06-group-integration-conos-analysis_cache/
Ignored: analysis/organoid-07-conos-visualization_cache/
Ignored: analysis/organoid-07-group-integration-conos-visualization_cache/
Ignored: analysis/organoid-08-conos-comparison_cache/
Ignored: analysis/organoid-0x-sample_integration_cache/
Ignored: analysis/sample5_QC_cache/
Ignored: analysis/timepoints-01-organoid-integration_cache/
Ignored: analysis/timepoints-02-cluster-analysis_cache/
Ignored: data/.DS_Store
Ignored: data/._.DS_Store
Ignored: data/._.smbdeleteAAA17ed8b4b
Ignored: data/._Lam_figure2_markers.R
Ignored: data/._README.md
Ignored: data/._Reactive_astrocytes_markers.xlsx
Ignored: data/._known_NSC_markers.R
Ignored: data/._known_cell_type_markers.R
Ignored: data/._metadata.csv
Ignored: data/._virus_cell_tropism_markers.R
Ignored: data/._~$Reactive_astrocytes_markers.xlsx
Ignored: data/data_sushi/
Ignored: data/filtered_feature_matrices/
Ignored: output/.DS_Store
Ignored: output/._.DS_Store
Ignored: output/._NSC_cluster2_marker_genes.txt
Ignored: output/._TDP-06-no_integration_cluster12_marker_genes.txt
Ignored: output/._TDP-06-no_integration_cluster13_marker_genes.txt
Ignored: output/._organoid_integration_cluster1_marker_genes.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_0.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_1.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_10.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_11.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_12.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_13.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_14.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_5.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_7.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_8.txt
Ignored: output/._tbl_TDP-08-01-muscat_cluster_all.xlsx
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_0.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_1.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_10.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_11.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_12.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_13.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_14.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_5.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_7.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_8.txt
Ignored: output/._tbl_TDP-08-02-targets_hek_cluster_all.xlsx
Ignored: output/._~$tbl_TDP-08-02-targets_hek_cluster_all.xlsx
Ignored: output/CH-test-01-preprocessing.rds
Ignored: output/CH-test-01-preprocessing_singlets.rds
Ignored: output/CH-test-01-preprocessing_singlets_filtered.rds
Ignored: output/CH-test-01-preprocessing_so.rds
Ignored: output/CH-test-01-preprocessing_so_filtered.rds
Ignored: output/Lam-01-clustering.rds
Ignored: output/NSC_1_clustering.rds
Ignored: output/NSC_cluster1_marker_genes.txt
Ignored: output/NSC_cluster2_marker_genes.txt
Ignored: output/NSC_cluster3_marker_genes.txt
Ignored: output/NSC_cluster4_marker_genes.txt
Ignored: output/NSC_cluster5_marker_genes.txt
Ignored: output/NSC_cluster6_marker_genes.txt
Ignored: output/NSC_cluster7_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster0_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster10_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster11_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster12_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster13_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster14_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster15_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster16_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster17_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster1_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster2_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster3_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster4_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster5_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster6_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster7_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster8_marker_genes.txt
Ignored: output/TDP-06-no_integration_cluster9_marker_genes.txt
Ignored: output/TDP-06_scran_markers.rds
Ignored: output/additional_filtering.rds
Ignored: output/conos/
Ignored: output/conos_organoid-06-conos-analysis.rds
Ignored: output/conos_organoid-06-group-integration-conos-analysis.rds
Ignored: output/figures/
Ignored: output/organoid_integration_cluster10_marker_genes.txt
Ignored: output/organoid_integration_cluster11_marker_genes.txt
Ignored: output/organoid_integration_cluster12_marker_genes.txt
Ignored: output/organoid_integration_cluster13_marker_genes.txt
Ignored: output/organoid_integration_cluster14_marker_genes.txt
Ignored: output/organoid_integration_cluster15_marker_genes.txt
Ignored: output/organoid_integration_cluster16_marker_genes.txt
Ignored: output/organoid_integration_cluster17_marker_genes.txt
Ignored: output/organoid_integration_cluster1_marker_genes.txt
Ignored: output/organoid_integration_cluster2_marker_genes.txt
Ignored: output/organoid_integration_cluster3_marker_genes.txt
Ignored: output/organoid_integration_cluster4_marker_genes.txt
Ignored: output/organoid_integration_cluster5_marker_genes.txt
Ignored: output/organoid_integration_cluster6_marker_genes.txt
Ignored: output/organoid_integration_cluster7_marker_genes.txt
Ignored: output/organoid_integration_cluster8_marker_genes.txt
Ignored: output/organoid_integration_cluster9_marker_genes.txt
Ignored: output/res_TDP-08-01-muscat.rds
Ignored: output/sce_01_preprocessing.rds
Ignored: output/sce_02_quality_control.rds
Ignored: output/sce_03_filtering.rds
Ignored: output/sce_03_filtering_all_genes.rds
Ignored: output/sce_06-1-prepare-sce.rds
Ignored: output/sce_TDP-08-01-muscat.rds
Ignored: output/sce_TDP_01_preprocessing.rds
Ignored: output/sce_TDP_02_quality_control.rds
Ignored: output/sce_TDP_03_filtering.rds
Ignored: output/sce_TDP_03_filtering_all_genes.rds
Ignored: output/sce_organoid-01-clustering.rds
Ignored: output/sce_preprocessing.rds
Ignored: output/so_04-stage_integration.rds
Ignored: output/so_04_1_cell_cycle.rds
Ignored: output/so_04_clustering.rds
Ignored: output/so_06-clustering_all_timepoints.rds
Ignored: output/so_08-00_clustering_HA_D96.rds
Ignored: output/so_08-clustering_timeline_HA.rds
Ignored: output/so_0x-sample_integration.rds
Ignored: output/so_TDP-06-cluster-analysis.rds
Ignored: output/so_TDP_04_clustering.rds
Ignored: output/so_TDP_05_plasmid_expression.rds
Ignored: output/so_additional_filtering_clustering.rds
Ignored: output/so_integrated_organoid-02-integration.rds
Ignored: output/so_merged_organoid-02-integration.rds
Ignored: output/so_organoid-01-clustering.rds
Ignored: output/so_sample_organoid-01-clustering.rds
Ignored: output/so_timepoints-01-organoid_integration.rds
Ignored: output/tbl_TDP-08-01-muscat.rds
Ignored: output/tbl_TDP-08-01-muscat_cluster_0.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_1.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_10.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_11.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_12.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_13.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_14.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_5.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_7.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_8.txt
Ignored: output/tbl_TDP-08-01-muscat_cluster_all.xlsx
Ignored: output/tbl_TDP-08-02-targets_hek.rds
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_0.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_1.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_10.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_11.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_12.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_13.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_14.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_5.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_7.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_8.txt
Ignored: output/tbl_TDP-08-02-targets_hek_cluster_all.xlsx
Ignored: output/~$tbl_TDP-08-02-targets_hek_cluster_all.xlsx
Ignored: scripts/.DS_Store
Ignored: scripts/._.DS_Store
Ignored: scripts/._bu_Rcode.R
Ignored: scripts/._plasmid_expression.sh
Ignored: scripts/._plasmid_expression_cell_hashing_test.sh
Ignored: scripts/._prepare_salmon_transcripts.R
Untracked files:
Untracked: Filtered.pdf
Untracked: Rplots.pdf
Untracked: Unfiltered
Untracked: Unfiltered.pdf
Untracked: analysis/CH-test-03-cluster-analysis.Rmd
Untracked: analysis/Lam-0-NSC_no_integration.Rmd
Untracked: analysis/TDP-07-01-STMN2_expression copy.Rmd
Untracked: analysis/additional_filtering.Rmd
Untracked: analysis/additional_filtering_clustering.Rmd
Untracked: analysis/organoid-01-1-qualtiy-control.Rmd
Untracked: analysis/organoid-06-conos-analysis-Seurat.Rmd
Untracked: analysis/organoid-06-conos-analysis-function.Rmd
Untracked: analysis/organoid-07-conos-visualization.Rmd
Untracked: analysis/organoid-07-group-integration-conos-visualization.Rmd
Untracked: analysis/organoid-08-conos-comparison.Rmd
Untracked: analysis/organoid-0x-sample_integration.Rmd
Untracked: analysis/sample5_QC.Rmd
Untracked: coverage.pdf
Untracked: coverage_sashimi.pdf
Untracked: coverage_sashimi.png
Untracked: data/Homo_sapiens.GRCh38.98.sorted.gtf
Untracked: data/Kanton_et_al/
Untracked: data/Lam_et_al/
Untracked: data/Sep2020/
Untracked: data/cell_hashing_test/
Untracked: data/cell_hashing_test_FB/
Untracked: data/reference/
Untracked: data/virus_cell_tropism_markers.R
Untracked: data/~$Reactive_astrocytes_markers.xlsx
Untracked: pbDS_cell_level.pdf
Untracked: pbDS_heatmap.pdf
Untracked: pbDS_top_expr_umap.pdf
Untracked: pbDS_upset.pdf
Untracked: sashimi.pdf
Untracked: scripts/bu_Rcode.R
Untracked: scripts/bu_code.Rmd
Untracked: scripts/plasmid_expression_cell_hashing_test.sh
Untracked: scripts/prepare_salmon_transcripts_cell_hashing_test.R
Untracked: scripts/salmon-latest_linux_x86_64/
Untracked: stmn2.pdf
Untracked: tdp.pdf
Unstaged changes:
Modified: analysis/05-annotation.Rmd
Modified: analysis/TDP-04-clustering.Rmd
Modified: analysis/TDP-08-01-HA-D96-expression-changes.Rmd
Modified: analysis/_site.yml
Modified: analysis/organoid-02-integration.Rmd
Modified: analysis/organoid-04-group_integration.Rmd
Modified: analysis/organoid-06-conos-analysis.Rmd
Note that any generated files, e.g. HTML, png, CSS, etc., are not included in this status report because it is ok for generated content to have uncommitted changes.
These are the previous versions of the repository in which changes were made to the R Markdown (analysis/CH-test-02-transgene-expression.Rmd) and HTML (docs/CH-test-02-transgene-expression.html) files. If you've configured a remote Git repository (see ?wflow_git_remote), click on the hyperlinks in the table below to view the files as they were in that past version.
| File | Version | Author | Date | Message |
|---|---|---|---|---|
| Rmd | e616506 | khembach | 2021-05-19 | add transgenes quantification and run clustering |
Load packages
library(cowplot)
library(ggplot2)
library(Seurat)
library(SingleCellExperiment)
library(tximport)
library(readxl)
library(dplyr)Load data
We combine the quantification of the plasmid transcript and the endogenous TDP-43 with the CellRanger count matrix.
meta <- read_excel(file.path("data", "cell_hashing_test", "SampleName_feature_ref_MHP.xlsm"))
so1 <- readRDS(file.path("output", "CH-test-01-preprocessing_singlets.rds"))# import alevin quants
txi <- tximport(file.path("data" , "cell_hashing_test", "alevin_STMN2",
"HashTag_test", "alevin", "quants_mat.gz"),
type="alevin")$countsWarning in Matrix::sparseMatrix(i = unlist(gene.idx), j = cell.idx, x =
counts.vec, : 'giveCsparse' has been deprecated; setting 'repr = "T"' for you## match the alevin and CellRanger cell IDs
colnames(txi) <- paste0(colnames(txi), "-1.cell_hashing_test")
rownames(txi) <- c("ENSG00000120948.TARDBP-alevin", "TDP43-HA", "GA50-EGFP",
"HA-GA50", "ENSG00000104435.STMN2-alevin")Add alevin counts to the Seurat object
rnames <- rownames(so1@assays$RNA@counts)
## add alevin counts to RNA matrix
counts <- rbind(so1@assays$RNA@counts,
as(matrix(0, nrow = nrow(txi), ncol = ncol(so1)),
"dgCMatrix"))
rownames(counts) <- c(rnames, rownames(txi))
## match the barcodes
m <- match(colnames(so1), colnames(txi))
counts[rownames(txi), which(!is.na(m))] <- txi[,m[!is.na(m)]]
so <- CreateSeuratObject(
counts = counts,
meta.data = so1@meta.data,
project = "cell_hashing_test")Label cells with demultiplexed sample
We use the HTO classification to define the original sample IDs.
so$HTO_classification %>% table.
B0253 B0254 B0257
1424 1329 1699 so$sample_id <- meta$Transgene[match(so$HTO_classification, meta$id)]
so$sample_id %>% table.
GA50-EGFP HA-GA50 TDP-43-HA
1699 1329 1424 Normalization
## log normalize the data using a scaling factor of 10000
so <- NormalizeData(so, verbose = FALSE, scale.factor = 10000,
normalization.method = "LogNormalize")so <- FindVariableFeatures(so, nfeatures = 2000,
selection.method = "vst", verbose = FALSE)
so <- ScaleData(so, verbose = FALSE, vars.to.regress = c("sum",
"subsets_Mt_percent"))Dimension reduction
We perform dimension reduction with t-SNE and UMAP based on PCA results.
so <- RunPCA(so, npcs = 30, verbose = FALSE)
so <- RunTSNE(so, reduction = "pca", dims = seq_len(20),
seed.use = 1, do.fast = TRUE, verbose = FALSE)
so <- RunUMAP(so, reduction = "pca", dims = seq_len(20),
seed.use = 1, verbose = FALSE)Plot PCA results
# top genes that are associated with the first two PCs
VizDimLoadings(so, dims = 1:2, reduction = "pca")
## PCA plot
DimPlot(so, reduction = "pca", group.by = "sample_id")
# elbow plot with the ranking of PCs based on the % of variance explained
ElbowPlot(so, ndims = 30)
Clustering
We cluster the cells using the reduced PCA dimensions.
so <- FindNeighbors(so, reduction = "pca", dims = seq_len(20), verbose = FALSE)
for (res in c(0.2, 0.4, 0.8, 1))
so <- FindClusters(so, resolution = res, random.seed = 1, verbose = FALSE)Dimension reduction plots
We plot the dimension reduction (DR) and color by sample, group and cluster ID
thm <- theme(aspect.ratio = 1, legend.position = "none")
ps <- lapply(c("sample_id", "ident"), function(u) {
p1 <- DimPlot(so, reduction = "tsne", group.by = u) + thm
p2 <- DimPlot(so, reduction = "umap", group.by = u)
lgd <- get_legend(p2)
p2 <- p2 + thm
list(p1, p2, lgd)
plot_grid(p1, p2, lgd, nrow = 1,
rel_widths = c(1, 1, 0.5))
})
plot_grid(plotlist = ps, ncol = 1)
QC on DR plots
.plot_features <- function(so, dr, id) {
FeaturePlot(so, features = id, reduction = dr, pt.size = 0.4,
cols = c("grey", "blue")) +
guides(col = guide_colourbar()) +
theme_void() + theme(aspect.ratio = 1)
}
ids <- c("sum", "detected", "subsets_Mt_percent", "ENSG00000120948.TARDBP",
"ENSG00000120948.TARDBP-alevin", "TDP43-HA", "GA50-EGFP", "HA-GA50",
"ENSG00000104435.STMN2-alevin", "ENSG00000104435.STMN2")
for (id in ids) {
cat("### ", id, "\n")
p1 <- .plot_features(so, "tsne", id)
lgd <- get_legend(p1)
p1 <- p1 + theme(legend.position = "none") + ggtitle("tSNE")
p2 <- .plot_features(so, "umap", id) + theme(legend.position = "none") +
ggtitle("UMAP")
ps <- plot_grid(plotlist = list(p1, p2), nrow = 1)
p <- plot_grid(ps, lgd, nrow = 1, rel_widths = c(1, 0.2))
print(p)
cat("\n\n")
}sum
detected
subsets_Mt_percent
ENSG00000120948.TARDBP
ENSG00000120948.TARDBP-alevin
TDP43-HA
GA50-EGFP
HA-GA50
ENSG00000104435.STMN2-alevin
ENSG00000104435.STMN2
Save Seurat object to RDS
saveRDS(so, file.path("output", "so_CH-test-02-transgene_expression.rds"))
sessionInfo()R version 4.0.5 (2021-03-31)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Ubuntu 18.04.5 LTS
Matrix products: default
BLAS: /usr/local/R/R-4.0.5/lib/libRblas.so
LAPACK: /usr/local/R/R-4.0.5/lib/libRlapack.so
locale:
[1] LC_CTYPE=en_US.UTF-8 LC_NUMERIC=C
[3] LC_TIME=en_US.UTF-8 LC_COLLATE=en_US.UTF-8
[5] LC_MONETARY=en_US.UTF-8 LC_MESSAGES=en_US.UTF-8
[7] LC_PAPER=en_US.UTF-8 LC_NAME=C
[9] LC_ADDRESS=C LC_TELEPHONE=C
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
attached base packages:
[1] parallel stats4 stats graphics grDevices utils datasets
[8] methods base
other attached packages:
[1] dplyr_1.0.2 readxl_1.3.1
[3] tximport_1.16.1 SingleCellExperiment_1.10.1
[5] SummarizedExperiment_1.18.1 DelayedArray_0.14.0
[7] matrixStats_0.56.0 Biobase_2.48.0
[9] GenomicRanges_1.40.0 GenomeInfoDb_1.24.2
[11] IRanges_2.22.2 S4Vectors_0.26.1
[13] BiocGenerics_0.34.0 SeuratObject_4.0.1
[15] Seurat_4.0.1 ggplot2_3.3.2
[17] cowplot_1.0.0 workflowr_1.6.2
loaded via a namespace (and not attached):
[1] Rtsne_0.15 colorspace_1.4-1 deldir_0.2-10
[4] ellipsis_0.3.1 ggridges_0.5.2 rprojroot_1.3-2
[7] XVector_0.28.0 fs_1.5.0 spatstat.data_2.1-0
[10] farver_2.0.3 leiden_0.3.3 listenv_0.8.0
[13] ggrepel_0.8.2 RSpectra_0.16-0 codetools_0.2-16
[16] splines_4.0.5 knitr_1.29 polyclip_1.10-0
[19] jsonlite_1.7.2 ica_1.0-2 cluster_2.1.0
[22] png_0.1-7 uwot_0.1.10 shiny_1.5.0
[25] sctransform_0.3.2 spatstat.sparse_2.0-0 compiler_4.0.5
[28] httr_1.4.2 backports_1.1.9 Matrix_1.3-3
[31] fastmap_1.0.1 lazyeval_0.2.2 later_1.1.0.1
[34] htmltools_0.5.0 tools_4.0.5 igraph_1.2.5
[37] GenomeInfoDbData_1.2.3 gtable_0.3.0 glue_1.4.2
[40] RANN_2.6.1 reshape2_1.4.4 rappdirs_0.3.1
[43] Rcpp_1.0.5 scattermore_0.7 cellranger_1.1.0
[46] vctrs_0.3.4 nlme_3.1-148 lmtest_0.9-37
[49] xfun_0.15 stringr_1.4.0 globals_0.12.5
[52] mime_0.9 miniUI_0.1.1.1 lifecycle_1.0.0
[55] irlba_2.3.3 goftest_1.2-2 future_1.17.0
[58] zlibbioc_1.34.0 MASS_7.3-51.6 zoo_1.8-8
[61] scales_1.1.1 spatstat.core_2.1-2 promises_1.1.1
[64] spatstat.utils_2.1-0 RColorBrewer_1.1-2 yaml_2.2.1
[67] reticulate_1.16 pbapply_1.4-2 gridExtra_2.3
[70] rpart_4.1-15 stringi_1.4.6 bitops_1.0-6
[73] rlang_0.4.10 pkgconfig_2.0.3 evaluate_0.14
[76] lattice_0.20-41 ROCR_1.0-11 purrr_0.3.4
[79] tensor_1.5 labeling_0.3 patchwork_1.0.1
[82] htmlwidgets_1.5.1 tidyselect_1.1.0 RcppAnnoy_0.0.18
[85] plyr_1.8.6 magrittr_1.5 R6_2.4.1
[88] generics_0.0.2 pillar_1.4.6 whisker_0.4
[91] withr_2.4.1 mgcv_1.8-31 fitdistrplus_1.1-1
[94] RCurl_1.98-1.3 survival_3.2-3 abind_1.4-5
[97] tibble_3.0.3 future.apply_1.6.0 crayon_1.3.4
[100] KernSmooth_2.23-17 spatstat.geom_2.1-0 plotly_4.9.2.1
[103] rmarkdown_2.3 grid_4.0.5 data.table_1.12.8
[106] git2r_0.27.1 digest_0.6.25 xtable_1.8-4
[109] tidyr_1.1.0 httpuv_1.5.4 munsell_0.5.0
[112] viridisLite_0.3.0