From bf2f099558cb7e2f43ae2cc360f17c74b56a9e07 Mon Sep 17 00:00:00 2001
From: Patrick Deelen <patrickdeelen@gmail.com>
Date: Sun, 13 Nov 2022 22:09:51 +0100
Subject: [PATCH] Update normalizeRecount3PerTissueAndDoPca.R

---
 .../normalizeRecount3PerTissueAndDoPca.R      | 61 ++++++++++++++++---
 1 file changed, 51 insertions(+), 10 deletions(-)

diff --git a/Downstreamer/src/main/r/downstreamer_main/recount3/normalizeRecount3PerTissueAndDoPca.R b/Downstreamer/src/main/r/downstreamer_main/recount3/normalizeRecount3PerTissueAndDoPca.R
index 18e52fac2..b0e3d8f20 100644
--- a/Downstreamer/src/main/r/downstreamer_main/recount3/normalizeRecount3PerTissueAndDoPca.R
+++ b/Downstreamer/src/main/r/downstreamer_main/recount3/normalizeRecount3PerTissueAndDoPca.R
@@ -4,7 +4,7 @@
 
 
 
-remoter::client("localhost", port = 55508)
+remoter::client("localhost", port = 55506)
 
 library(DESeq2)
 library(parallel)
@@ -65,6 +65,50 @@ perTissueExp <- lapply(tissueClasses, function(tissue){
 #names(perTissueExp) <- tissueClasses
 #save(perTissueExp, file = "perTissueNormalization/selectedSamplesRawExpressionPerTissue.RData")
 tissue = "Kidney"
+load(file = "Metadata/combinedMeta_2022_09_15.RData", verbose = T)
+
+covariatesToCorrectFor <- read.delim("CovariateNames.txt", header = F)$V1
+
+str(sum(is.na(combinedMeta[,"sra.sample_spots"])))
+
+#TCGA samples don't have sra.sample_spots instead use recount_qc.bc_frag.count
+missingSampleSpots <- is.na(combinedMeta[,"sra.sample_spots"])
+combinedMeta[missingSampleSpots,"sra.sample_spots"] <- combinedMeta[missingSampleSpots,"recount_qc.bc_frag.count"]
+
+#TCGA and Gtex don't report layout but is all paired
+missingLayout <- is.na(combinedMeta[,"sra.library_layout"])
+combinedMeta[missingLayout,"sra.library_layout"] <- "paired"
+
+combinedMetaSelection <- combinedMeta[,covariatesToCorrectFor]
+combinedMetaSelection$sra.library_layout <- as.factor(combinedMetaSelection$sra.library_layout)
+table(combinedMeta[,"sra.library_layout"],useNA = "a")
+
+cl <- makeCluster(20)
+
+tissue <- "Salivary Gland-Minor Salivary Gland"
+
+sink <- lapply(tissueClasses, function(tissue){
+  cat(tissue, "\n")
+  
+  
+  load(file = paste0("perTissueNormalization/vstExp/",make.names(tissue),".RData"))
+
+  combinedMetaTissue <- combinedMetaSelection[colnames(vstExp),]
+  if(min(table(combinedMetaTissue$sra.library_layout)) < 10){
+    combinedMetaTissue <- combinedMetaTissue[,-match("sra.library_layout",colnames(combinedMetaTissue))]
+  }
+  
+  vstExpCovCor <- parApply(cl, vstExp, 1 ,function(geneExp, combinedMetaTissue){
+    return(residuals(lm(geneExp ~ . ,data = combinedMetaTissue)))
+  }, combinedMetaTissue = combinedMetaTissue)
+  vstExpCovCor <- t(vstExpCovCor)
+  
+  
+  save(vstExpCovCor, file = paste0("perTissueNormalization/vstExpCovCor/",make.names(tissue),".RData"))
+
+})
+
+stopCluster(cl)
 
 
 sink <- lapply(tissueClasses, function(tissue){
@@ -74,7 +118,7 @@ sink <- lapply(tissueClasses, function(tissue){
   load(file = paste0("perTissueNormalization/vstExp/",make.names(tissue),".RData"))
   
   #https://stackoverflow.com/questions/18964837/fast-correlation-in-r-using-c-and-parallelization/18965892#18965892
-  expScale = vstExp - rowMeans(vstExp);
+  expScale = vstExpCovCor - rowMeans(vstExpCovCor);
   # Standardize each variable
     expScale = expScale / sqrt(rowSums(expScale^2));   
   #expCov = tcrossprod(expScale);#equevelent to correlation due to center scale
@@ -121,7 +165,6 @@ eigenVec <- tissueVstPca$eigenVectors
 rownames(vstExp) <- (gsub("\\..+", "", rownames(eigenVec)))
 write.table(eigenVec, file = gzfile(paste0("/groups/umcg-fg/tmp01/projects/genenetwork/recount3/perTissueNormalization/vstPca/",make.names(tissue),"_eigenVec.txt.gz")), sep = "\t", col.names = NA, quote = F)
 
-load(file = "Metadata/combinedMeta_2022_09_15.RData", verbose = T)
 
 metaNumeric <- as.matrix(combinedMeta[,sapply(combinedMeta, is.numeric)])
 
@@ -134,7 +177,7 @@ marCorZPerTissue <- lapply(tissueClasses, function(tissue){
   pcs <- tissueVstPca$expPcs
   metaTest <- metaNumeric[rownames(pcs),]
 
-  metaVPcsZ <- apply(expPcs, 2, function(x){
+  metaVPcsZ <- apply(pcs, 2, function(x){
     apply(metaTest, 2, function(y, x){
       z <- !is.na(x) & !is.na(y)
       if(sum(z) < 10){
@@ -157,7 +200,7 @@ str(marCorZPerTissue)
 marCorZPerTissue2 <- do.call(cbind, marCorZPerTissue)
 str(marCorZPerTissue2)
 
-sort(apply(marCorZPerTissue2,1,max))
+covariateZscores <- apply(marCorZPerTissue2,1,mean)
 
 marCorZPerTissue2["recount_qc.star.%_of_reads_mapped_to_multiple_loci",]
 library(vioplot)
@@ -174,6 +217,8 @@ cor.test(pcs[,1], metaTest[,"recount_qc.star.%_of_reads_mapped_to_multiple_loci"
 plot(pcs[,1], metaTest[,"recount_qc.star.%_of_reads_mapped_to_multiple_loci"], pch = 16, col=adjustcolor("grey", alpha.f = 0.5))
 dev.off()
 
+pcs <- expPcs
+
 
 tissueSamplesInfo <- samplesWithPredictionNoOutliers[rownames(pcs),]
 studies <- length(unique(tissueSamplesInfo$study))
@@ -184,7 +229,7 @@ rpng(width = 1500, height = 1500)
 palette(adjustcolor(viridis(studies, option = "H"), alpha.f = 0.5))
 pchMap <- rep(c(15,16,17), length.out = studies)
 plot(pcs[,1],metaTest[,"recount_qc.star.%_of_reads_mapped_to_multiple_loci"], col = as.factor(tissueSamplesInfo$study), pch = pchMap[as.factor(tissueSamplesInfo$study)], cex = 2, main = paste0("Studies (", studies,")"), xlab = paste0("Comp 1 (", round(explainedVariance[1],2) ,"%)"), ylab = "Percentage read map multiple loci", bty = "n")
-
+plot(pcs[,1],pcs[,2], col = as.factor(tissueSamplesInfo$study), pch = pchMap[as.factor(tissueSamplesInfo$study)], cex = 2, main = paste0("Studies (", studies,")"), xlab = paste0("Comp 1 (", round(explainedVariance[1],2) ,"%)"), ylab = "Percentage read map multiple loci", bty = "n")
 dev.off()
 
 
@@ -202,10 +247,6 @@ dev.off()
 
 
 
-
-
-
-
 #OLD