## ----include = FALSE---------------------------------------------------------- knitr::opts_chunk$set( collapse = FALSE ) ## ----setup, message=FALSE, warning=FALSE-------------------------------------- library(miloR) library(SingleCellExperiment) library(scater) library(scran) library(dplyr) library(patchwork) ## ----------------------------------------------------------------------------- data("sim_trajectory", package = "miloR") ## Extract SingleCellExperiment object traj_sce <- sim_trajectory[['SCE']] ## Extract sample metadata to use for testing traj_meta <- sim_trajectory[["meta"]] ## Add metadata to colData slot colData(traj_sce) <- DataFrame(traj_meta) ## ----------------------------------------------------------------------------- logcounts(traj_sce) <- log(counts(traj_sce) + 1) traj_sce <- runPCA(traj_sce, ncomponents=30) traj_sce <- runUMAP(traj_sce) plotUMAP(traj_sce) ## ----------------------------------------------------------------------------- traj_milo <- Milo(traj_sce) reducedDim(traj_milo, "UMAP") <- reducedDim(traj_sce, "UMAP") traj_milo ## ----eval=FALSE--------------------------------------------------------------- # library(zellkonverter) # # # Obtaining an example H5AD file. # example_h5ad <- system.file("extdata", "krumsiek11.h5ad", # package = "zellkonverter") # # example_h5ad_sce <- readH5AD(example_h5ad) # example_h5ad_milo <- Milo(example_h5ad_sce) ## ----eval=FALSE--------------------------------------------------------------- # library(Seurat) # data("pbmc_small") # pbmc_small_sce <- as.SingleCellExperiment(pbmc_small) # pbmc_small_milo <- Milo(pbmc_small_sce) ## ----------------------------------------------------------------------------- traj_milo <- buildGraph(traj_milo, k = 10, d = 30) ## ----------------------------------------------------------------------------- traj_milo <- makeNhoods(traj_milo, prop = 0.1, k = 10, d=30, refined = TRUE) ## ----------------------------------------------------------------------------- plotNhoodSizeHist(traj_milo) ## ----------------------------------------------------------------------------- traj_milo <- countCells(traj_milo, meta.data = data.frame(colData(traj_milo)), samples="Sample") ## ----------------------------------------------------------------------------- head(nhoodCounts(traj_milo)) ## ----------------------------------------------------------------------------- traj_design <- data.frame(colData(traj_milo))[,c("Sample", "Condition")] traj_design <- distinct(traj_design) rownames(traj_design) <- traj_design$Sample ## Reorder rownames to match columns of nhoodCounts(milo) traj_design <- traj_design[colnames(nhoodCounts(traj_milo)), , drop=FALSE] traj_design ## ----------------------------------------------------------------------------- traj_milo <- calcNhoodDistance(traj_milo, d=30) ## ----------------------------------------------------------------------------- rownames(traj_design) <- traj_design$Sample da_results <- testNhoods(traj_milo, design = ~ Condition, design.df = traj_design) ## ----------------------------------------------------------------------------- da_results %>% arrange(- SpatialFDR) %>% head() ## ----fig.width=10, fig.height=6----------------------------------------------- traj_milo <- buildNhoodGraph(traj_milo) plotUMAP(traj_milo) + plotNhoodGraphDA(traj_milo, da_results, alpha=0.05) + plot_layout(guides="collect") ## ----------------------------------------------------------------------------- sessionInfo()