Examine B-cell and NK cell topics and clusters
Peter Carbonetto
Last updated: 2020-09-08
Checks: 6 1
Knit directory: single-cell-topics/analysis/
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- diff-count-analysis-clusters-68k
- diff-count-analysis-clusters-purified
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| File | Version | Author | Date | Message |
|---|---|---|---|---|
| Rmd | 4525eda | Peter Carbonetto | 2020-09-08 | workflowr::wflow_publish(“bcells_and_nkcells.Rmd”) |
| html | 2e5917d | Peter Carbonetto | 2020-09-08 | Performed first build of bcells_and_nkcells page. |
| Rmd | faea79b | Peter Carbonetto | 2020-09-08 | workflowr::wflow_publish(“bcells_and_nkcells.Rmd”) |
| Rmd | 681efe3 | Peter Carbonetto | 2020-09-08 | Added steps to bcells_and_nkcells analysis to run cluster-based differential expression analysis. |
| Rmd | 77499e2 | Peter Carbonetto | 2020-09-07 | Implemented first steps of bcells_and_nkcells analysis. |
| Rmd | d749c17 | Peter Carbonetto | 2020-09-07 | Added notes to plots_pbmc; committed pbmc-clustering.RData output. |
Add text here.
Load the packages used in the analysis below.
library(Matrix)
library(fastTopics)
library(ggplot2)
library(ggrepel)
library(cowplot)
source("../code/more_plots.R")Load the mixture of FACS-purified PBMC data, the \(k = 6\) Poisson NMF model fit, and the results of the differential expression analysis using this model fit.
fit_purified <-
readRDS("../output/pbmc-purified/rds/fit-pbmc-purified-scd-ex-k=6.rds")$fit
load("../output/pbmc-purified/postfit-pbmc-purified-scd-ex-k=6.RData")
load("../data/pbmc_purified.RData")
ids <- rownames(diff_count_res$Z)
counts_purified <- counts[,ids]
genes_purified <- genes[match(ids,genes$ensembl),]
diff_count_purified <- diff_count_res
rm(samples,genes,counts,diff_count_res,gene_info,ids)
rm(gene_set_info,gene_sets,gsea_res)Load the “unsorted” 68k PBMC data, the \(k = 6\) Poisson NMF model fit, and the results of the differential expression analysis using this model fit.
fit_68k <- readRDS("../output/pbmc-68k/rds/fit-pbmc-68k-scd-ex-k=6.rds")$fit
load("../output/pbmc-68k/postfit-pbmc-68k-scd-ex-k=6.RData")
load("../data/pbmc_68k.RData")
ids <- rownames(diff_count_res$Z)
counts_68k <- counts[,ids]
genes_68k <- genes[match(ids,genes$ensembl),]
diff_count_68k <- diff_count_res
rm(samples,genes,counts,diff_count_res,ids,gene_set_info,gene_sets,gsea_res)Load the clustering of the purified and 68k data set that was determined in the “plots_pbmc” analysis.
load("../output/pbmc-clustering.RData")Perform a differential expression analysis using the clustering of the purified data.
fit_clusters_purified <-
init_poisson_nmf_from_clustering(counts_purified,samples_purified$cluster)
diff_count_clusters_purified <- diff_count_analysis(fit_clusters_purified,
counts_purified)
# All topic proportions are either zero or one; using simpler single-topic calculations for model parameter estimates
# Fitting 17316 x 6 = 103896 univariate Poisson models.
# Computing log-fold change statistics.
Warning: The above code chunk cached its results, but it won’t be re-run if previous chunks it depends on are updated. If you need to use caching, it is highly recommended to also set knitr::opts_chunk$set(autodep = TRUE) at the top of the file (in a chunk that is not cached). Alternatively, you can customize the option dependson for each individual chunk that is cached. Using either autodep or dependson will remove this warning. See the knitr cache options for more details.
Perform a differential expression analysis using the clustering of the 68k data.
fit_clusters_68k <- init_poisson_nmf_from_clustering(counts_68k,
samples_68k$cluster)
diff_count_clusters_68k <- diff_count_analysis(fit_clusters_68k,counts_68k)
# All topic proportions are either zero or one; using simpler single-topic calculations for model parameter estimates
# Fitting 16402 x 9 = 147618 univariate Poisson models.
# Computing log-fold change statistics.
Warning: The above code chunk cached its results, but it won’t be re-run if previous chunks it depends on are updated. If you need to use caching, it is highly recommended to also set knitr::opts_chunk$set(autodep = TRUE) at the top of the file (in a chunk that is not cached). Alternatively, you can customize the option dependson for each individual chunk that is cached. Using either autodep or dependson will remove this warning. See the knitr cache options for more details.
In the purified data set, compare the gene-wise z-scores from topic 3 to the \(z\)-scores from cluster B.
p1 <- diff_count_scatterplot(diff_count_clusters_purified$Z[,"B"],
diff_count_purified$Z[,3],
diff_count_purified$colmeans,
genes_purified$symbol,
label_above_score = 100) +
labs(x = "cluster B",y = "topic 3",title = "z-scores")
print(p1)
In the 68k data, compare the gene-wise z-scores from topic 5 to the \(z\)-scores from cluster A2.
p2 <- diff_count_scatterplot(diff_count_clusters_68k$Z[,"A2"],
diff_count_68k$Z[,5],
diff_count_68k$colmeans,
genes_68k$symbol,
label_above_score = 100) +
labs(x = "cluster A2",y = "topic 5",title = "z-scores")
print(p2)
| Version | Author | Date |
|---|---|---|
| 2e5917d | Peter Carbonetto | 2020-09-08 |
Plot histograms of the topic 3 proportions inside and outside cluster B in the purified data.
fit <- poisson2multinom(fit_purified)
p3 <- ggplot(data.frame(x = fit$L[,3]),aes(x)) +
geom_histogram(bins = 50,color = "white",fill = "black",na.rm = TRUE) +
ylim(c(0,2000)) +
labs(x = "topic 3 proportion",y = "number of cells") +
theme_cowplot(font_size = 10)
p4 <- ggplot(data.frame(x = fit$L[samples_purified$cluster == "B",3]),aes(x)) +
geom_histogram(bins = 30,color = "white",fill = "black",na.rm = TRUE) +
labs(x = "topic 3 proportion",y = "number of cells") +
theme_cowplot(font_size = 10)
plot_grid(p3,p4)
Plot histograms of the topic 5 proportions inside and outside cluster A3 in the 68k data.
fit <- poisson2multinom(fit_68k)
p5 <- ggplot(data.frame(x = fit$L[,5]),aes(x)) +
geom_histogram(bins = 50,color = "white",fill = "black",na.rm = TRUE) +
ylim(c(0,1200)) +
labs(x = "topic 5 proportion",y = "number of cells") +
theme_cowplot(font_size = 10)
p6 <- ggplot(data.frame(x = fit$L[samples_68k$cluster == "A2",5]),aes(x)) +
geom_histogram(bins = 30,color = "white",fill = "black",na.rm = TRUE) +
labs(x = "topic 5 proportion",y = "number of cells") +
theme_cowplot(font_size = 10)
plot_grid(p5,p6)
sessionInfo()
# R version 3.6.2 (2019-12-12)
# Platform: x86_64-apple-darwin15.6.0 (64-bit)
# Running under: macOS Catalina 10.15.6
#
# Matrix products: default
# BLAS: /Library/Frameworks/R.framework/Versions/3.6/Resources/lib/libRblas.0.dylib
# LAPACK: /Library/Frameworks/R.framework/Versions/3.6/Resources/lib/libRlapack.dylib
#
# locale:
# [1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8
#
# attached base packages:
# [1] stats graphics grDevices utils datasets methods base
#
# other attached packages:
# [1] cowplot_1.0.0 ggrepel_0.9.0 ggplot2_3.3.0 fastTopics_0.3-174
# [5] Matrix_1.2-18
#
# loaded via a namespace (and not attached):
# [1] Rcpp_1.0.5 lattice_0.20-38 tidyr_1.0.0
# [4] prettyunits_1.1.1 assertthat_0.2.1 zeallot_0.1.0
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# [43] later_1.0.0 MASS_7.3-51.4 grid_3.6.2
# [46] jsonlite_1.6 gtable_0.3.0 lifecycle_0.1.0
# [49] git2r_0.26.1 magrittr_1.5 scales_1.1.0
# [52] RcppParallel_4.4.2 stringi_1.4.3 farver_2.0.1
# [55] fs_1.3.1 promises_1.1.0 vctrs_0.2.1
# [58] tools_3.6.2 glue_1.3.1 purrr_0.3.3
# [61] hms_0.5.2 yaml_2.2.0 colorspace_1.4-1
# [64] plotly_4.9.2 knitr_1.26 quantreg_5.54
# [67] MCMCpack_1.4-5