Copy VAMC15x analysis to VAMC13x

r.scripts/sc-TissueMapper_RUN.PrFx_VAMC013.R

+gc()
+library(methods)
+library(optparse)
+library(Seurat)
+library(readr)
+library(fBasics)
+library(pastecs)
+library(qusage)
+library(RColorBrewer)
+library(monocle)
+library(dplyr)
+library(viridis)
+library(readxl)
+
+source("../r.scripts/sc-TissueMapper.R")
+
+#Create folder structure
+setwd("../")
+if (!dir.exists("./analysis")){
+  dir.create("./analysis")
+}
+if (!dir.exists("./analysis/qc")){
+  dir.create("./analysis/qc")
+}
+if (!dir.exists("./analysis/qc/cc")){
+  dir.create("./analysis/qc/cc")
+}
+if (!dir.exists("./analysis/tSNE")){
+  dir.create("./analysis/tSNE")
+}
+if (!dir.exists("./analysis/tSNE/pre.stress")){
+  dir.create("./analysis/tSNE/pre.stress")
+}
+if (!dir.exists("./analysis/pca")){
+  dir.create("./analysis/pca")
+}
+if (!dir.exists("./analysis/pca/stress")){
+  dir.create("./analysis/pca/stress")
+}
+if (!dir.exists("./analysis/violin")){
+  dir.create("./analysis/violin")
+}
+if (!dir.exists("./analysis/violin/stress")){
+  dir.create("./analysis/violin/stress")
+}
+if (!dir.exists("./analysis/table")){
+  dir.create("./analysis/table")
+}
+if (!dir.exists("./analysis/tSNE/post.stress")){
+  dir.create("./analysis/tSNE/post.stress")
+}
+if (!dir.exists("./analysis/cor")){
+  dir.create("./analysis/cor")
+}
+if (!dir.exists("./analysis/tSNE/lin")){
+  dir.create("./analysis/tSNE/lin")
+}
+if (!dir.exists("./analysis/tSNE/epi")){
+  dir.create("./analysis/tSNE/epi")
+}
+if (!dir.exists("./analysis/tSNE/st")){
+  dir.create("./analysis/tSNE/st")
+}
+if (!dir.exists("./analysis/tSNE/merge")){
+  dir.create("./analysis/tSNE/merge")
+}
+if (!dir.exists("./analysis/pca/ne")){
+  dir.create("./analysis/pca/ne")
+}
+if (!dir.exists("./analysis/tSNE/ne")){
+  dir.create("./analysis/tSNE/ne")
+}
+if (!dir.exists("./analysis/violin/ne")){
+  dir.create("./analysis/violin/ne")
+}
+if (!dir.exists("./analysis/tSNE/FINAL")){
+  dir.create("./analysis/tSNE/FINAL")
+}
+if (!dir.exists("./analysis/deg")){
+  dir.create("./analysis/deg")
+}
+if (!dir.exists("./analysis/cca")){
+  dir.create("./analysis/cca")
+}
+if (!dir.exists("./analysis/diy")){
+  dir.create("./analysis/diy")
+}
+if (!dir.exists("./analysis/pseudotime")){
+  dir.create("./analysis/pseudotime")
+}
+
+#Retrieve command-line options
+option_list=list(
+  make_option("--p",action="store",default="DPrF",type='character',help="Project Name"),
+  make_option("--g",action="store",default="ALL",type='character',help="Group To analyze"),
+  make_option("--lg",action="store",default=500,type='integer',help="Threshold for cells with minimum genes"),
+  make_option("--hg",action="store",default=3000,type='integer',help="Threshold for cells with maximum genes"),
+  make_option("--lm",action="store",default=0,type='numeric',help="Threshold for cells with minimum %mito genes"),
+  make_option("--hm",action="store",default=0.1,type='numeric',help="Threshold for cells with maximum %mito genes"),
+  make_option("--lx",action="store",default=0.2,type='numeric',help="x low threshold for hvg selection"),
+  make_option("--hx",action="store",default=5,type='numeric',help="x high threshold for hvg selection"),
+  make_option("--ly",action="store",default=1,type='numeric',help="y low threshold for hvg selection"),
+  make_option("--cc",action="store",default=TRUE,type='logical',help="Scale cell cycle?"),
+  make_option("--cca",action="store",default=50,type='integer',help="Number of CCAs to cacluate"),
+  make_option("--acca",action="store",default=30,type='integer',help="Number of CCAs to align"),
+  make_option("--pc",action="store",default=50,type='integer',help="Number of PCs to cacluate"),
+  make_option("--res.prestress",action="store",default=1,type='numeric',help="Resolution to cluster, pre-stress"),
+  make_option("--st",action="store",default=TRUE,type='logical',help="Remove stressed cells?"),
+  make_option("--stg",action="store",default="dws",type='character',help="Geneset to use for stress ID"),
+  make_option("--cut.stress",action="store",default=0.9,type='numeric',help="Cutoff for stress score"),
+  make_option("--res.poststress",action="store",default=0.5,type='numeric',help="Resolution to cluster, post-stress"),
+  make_option("--cut.ne",action="store",default=0.999,type='numeric',help="Cutoff for NE score")
+)
+opt=parse_args(OptionParser(option_list=option_list))
+rm(option_list)
+if (opt$lm==0){opt$lm=-Inf}
+
+sc10x <- scLoad("VAMC013PrRdF")
+
+if (opt$cc==TRUE){
+  results <- scCellCycle(sc10x)
+  sc10x <- results[[1]]
+  genes.s <- results[[2]]
+  genes.g2m <- results[[3]]
+  rm(results)
+} else {
+  genes.s=""
+  genes.g2m=""
+}
+
+results <- scQC(sc10x,lg=opt$lg,hg=opt$hg,lm=opt$lm,hm=opt$hm)
+sc10x <- results[[1]]
+counts.cell.raw <- results[[2]]
+counts.gene.raw <- results[[3]]
+counts.cell.filtered <- results[[4]]
+counts.gene.filtered <- results[[5]]
+rm(results)
+
+gc()
+if (opt$cc==TRUE){
+  sc10x <- ScaleData(object=sc10x,vars.to.regress=c("nUMI","percent.mito","S.Score","G2M.Score"),display.progress=FALSE,do.par=TRUE,num.cores=45)
+} else {
+  sc10x <- ScaleData(object=sc10x,vars.to.regress=c("nUMI","percent.mito"),display.progress=FALSE,do.par=TRUE,num.cores=45)
+}
+gc()
+
+results <- scPC(sc10x,lx=opt$lx,hx=opt$hx,ly=opt$ly,cc=opt$cc,pc=50,hpc=0.85,file="pre.stress",cca=FALSE)
+sc10x <- results[[1]]
+genes.hvg.prestress <- results[[2]]
+pc.use.prestress <- results[[3]]
+rm(results)
+
+sc10x <- scCluster(sc10x,pc.use=pc.use.prestress,res.use=opt$res.prestress,folder="pre.stress",red="pca")
+
+if (opt$st==TRUE){
+  results <- scStress(sc10x,stg=opt$stg,res.use=opt$res.prestress,cut=opt$cut.stress)
+  sc10x <- results[[1]]
+  counts.cell.filtered.stress <- results[[2]]
+  sc10x.Stress <- results[[3]]
+  rm(results)
+  
+  results <- scPC(sc10x,lx=opt$lx,hx=opt$hx,ly=opt$ly,cc=opt$cc,pc=50,hpc=0.85,file="post.stress",cca=FALSE)
+  sc10x <- results[[1]]
+  genes.hvg.poststress <- results[[2]]
+  pc.use.poststress <- results[[3]]
+  rm(results)
+  
+  sc10x <- scCluster(sc10x,pc.use=pc.use.poststress,res.use=opt$res.poststress,folder="post.stress",red="pca")
+}
+
+gene.set1 <- read_delim("./genesets/genes.deg.Epi.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "Epi"
+gene.set <- c(gene.set1)
+gene.set1 <- read_delim("./genesets/genes.deg.St.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "St"
+gene.set <- c(gene.set,gene.set1)
+gene.set1 <- read_delim("./genesets/genes.deg.BE.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "BE"
+gene.set <- c(gene.set,gene.set1)
+gene.set1 <- read_delim("./genesets/genes.deg.LE.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "LE"
+gene.set <- c(gene.set,gene.set1)
+gene.set1 <- read_delim("./genesets/genes.deg.OE1.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "OE_SCGB"
+gene.set <- c(gene.set,gene.set1)
+gene.set1 <- read_delim("./genesets/genes.deg.OE2.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "OE_KRT13"
+gene.set <- c(gene.set,gene.set1)
+gene.set1 <- read_delim("./genesets/genes.deg.Endo.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "Endo"
+gene.set <- c(gene.set,gene.set1)
+gene.set1 <- read_delim("./genesets/genes.deg.SM.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "SM"
+gene.set <- c(gene.set,gene.set1)
+gene.set1 <- read_delim("./genesets/genes.deg.Fib.csv",",",escape_double=FALSE,trim_ws=TRUE,col_names=TRUE)
+gene.set1 <- gene.set1[1]
+gene.set1 <- as.list(gene.set1)
+names(gene.set1) <- "Fib"
+gene.set <- c(gene.set,gene.set1)
+genes.leu <- read_excel("genesets/40425_2017_215_MOESM1_ESM.xlsx",sheet="S3. Candidate markers")
+leu <- as.list(unique(genes.leu[,2]))$Cell
+leu.l <- leu[-c(1,3,4,7:9,14,15,17:18,20:21,23:30)]
+genes.leu <- genes.leu[unlist(genes.leu[,2]) %in% unlist(leu),]
+genes.leu.l <- genes.leu[unlist(genes.leu[,2]) %in% unlist(leu.l),]
+gene.set1 <- split(genes.leu.l[,1], genes.leu.l[,2])
+gene.set1 <- lapply(gene.set1,unname)
+gene.set1 <- lapply(gene.set1,unlist)
+gene.set <- c(gene.set,gene.set1)
+rm(gene.set1)
+gc()
+min.all <- min(table(sc10x@meta.data[,paste0("res",opt$res.poststress)]))
+results <- scQuSAGE(sc10x,gs=gene.set,res.use=opt$res.poststress,ds=min.all,nm="Pop",folder="lin")
+sc10x <- results[[1]]
+results.cor.Lin <- results[[2]]
+results.clust.Lin.id <- results[[3]]
+rm(results)
+rm(gene.set)
+
+sc10x.Epi.NE <- scNE(sc10x.Epi,neg="dws",cut=opt$cut.ne)