Investigating whether Seasol responsive genes are linked to abiotic stress
Mark Ziemann
2021-02-01
Source codes: https://github.com/markziemann/tohidul_rnaseq
Background
Generate a heatmap of expression:
Genes from 0 hr AN/DP vs Water (h2o vs 80 de) fold change of =>1.5 and padj value <0.05.
Expression data mined from eFP browser here: http://bar.utoronto.ca/affydb/cgi-bin/affy_db_exprss_browser_in.cgi?pub=&dataset=atgenexp_stress
suppressPackageStartupMessages({
library("gplots")
})Load data
Here we load the data produced by earlier analysis We use the Kallisto transcript quantifications and summarise these to gene level.
de1 <- read.table("de1.tsv",header=TRUE,row.names=1,sep="\t",fill=TRUE)
de1_up <- subset(de1,log2FoldChange>1.5 & padj < 0.05)$GeneID
writeLines(de1_up,con="de1up.txt")Then I searched in the eFP browser. Here's a list of Genes Not Found in the Database:
At1g07985, At1g29100, At1g47395, At1g53610, At1g58320, At2g36255, At4g11393, At4g25433, At4g36850, At5g46610, At5g54165
# numeric data first
datfile="exprss_text_unsorted_25077.txt"
efp <- read.table(datfile, header=TRUE, row.names=1, sep="\t", comment.char="#")
genenames <- efp[,1,drop=FALSE]
efp[,1]=NULL
# sample info data next
dat <- readLines(datfile)
dat <- dat[grep('#',dat)]
datlist <- strsplit(dat,"\t")
sampleinfo <- as.data.frame(do.call(rbind, datlist))
sampleinfo[1:5,1:6]## V1 V2 V3 V4 V5 V6
## 1 #category Cold Stress Cold Stress Cold Stress Cold Stress
## 2 #age 1.02 1.02 1.02 1.02
## 3 #tissue Shoot Root Shoot Root
## 4 #control Yes Yes Yes Yes
## 5 #mutant Wt Wt Wt Wtsampleinfo$V2=NULL
rownames(sampleinfo) <- gsub("#","",sampleinfo$V1)
sampleinfo$V1=NULL
sampleinfo <- as.data.frame(t(sampleinfo))
str(sampleinfo)## 'data.frame': 236 obs. of 7 variables:
## $ category : chr "Cold Stress" "Cold Stress" "Cold Stress" "Cold Stress" ...
## $ age : chr "1.02" "1.02" "1.02" "1.02" ...
## $ tissue : chr "Shoot" "Root" "Shoot" "Root" ...
## $ control : chr "Yes" "Yes" "Yes" "Yes" ...
## $ mutant : chr "Wt" "Wt" "Wt" "Wt" ...
## $ timecourse: chr "0.5 h" "0.5 h" "1.0 h" "1.0 h" ...
## $ alias : chr "Col-0" "Col-0" "Col-0" "Col-0" ...Filtering
# Shoot
cols <- which(
sampleinfo$timecourse =="1.0 h" &
sampleinfo$control =="No" &
sampleinfo$tissue =="Shoot" &
sampleinfo$age =="1.02" )
shoot <- efp[,cols]
shootsampleinfo <- sampleinfo[cols,]
shootsampleinfo## category age tissue control mutant timecourse alias
## V17 Cold Stress 1.02 Shoot No Wt 1.0 h Col-0
## V41 Osmotic Stress 1.02 Shoot No Wt 1.0 h Col-0
## V65 Salt Stress 1.02 Shoot No Wt 1.0 h Col-0
## V93 Drought Stress 1.02 Shoot No Wt 1.0 h Col-0
## V117 Genotoxic Stress 1.02 Shoot No Wt 1.0 h Col-0
## V141 Oxidative Stress 1.02 Shoot No Wt 1.0 h Col-0
## V169 UV-B Stress 1.02 Shoot No Wt 1.0 h Col-0
## V197 Wounding Stress 1.02 Shoot No Wt 1.0 h Col-0
## V227 Heat Stress 1.02 Shoot No Wt 1.0 h Col-0colnames(shoot) <- gsub(" Stress","",shootsampleinfo$category)
# Root
cols <- which(
sampleinfo$timecourse =="1.0 h" &
sampleinfo$control =="No" &
sampleinfo$tissue =="Root" &
sampleinfo$age =="1.02" )
root <- efp[,cols]
rootsampleinfo <- sampleinfo[cols,]
rootsampleinfo## category age tissue control mutant timecourse alias
## V18 Cold Stress 1.02 Root No Wt 1.0 h Col-0
## V42 Osmotic Stress 1.02 Root No Wt 1.0 h Col-0
## V66 Salt Stress 1.02 Root No Wt 1.0 h Col-0
## V94 Drought Stress 1.02 Root No Wt 1.0 h Col-0
## V118 Genotoxic Stress 1.02 Root No Wt 1.0 h Col-0
## V142 Oxidative Stress 1.02 Root No Wt 1.0 h Col-0
## V170 UV-B Stress 1.02 Root No Wt 1.0 h Col-0
## V198 Wounding Stress 1.02 Root No Wt 1.0 h Col-0
## V228 Heat Stress 1.02 Root No Wt 1.0 h Col-0colnames(root) <- gsub(" Stress","",rootsampleinfo$category)
# osmoroot
cols <- which(
sampleinfo$control =="No" &
sampleinfo$category =="Osmotic Stress" &
sampleinfo$tissue =="Root" )
osmoroot <- efp[,cols]
osmorootsampleinfo <- sampleinfo[cols,]
osmorootsampleinfo## category age tissue control mutant timecourse alias
## V40 Osmotic Stress 1.02 Root No Wt 0.5 h Col-0
## V42 Osmotic Stress 1.02 Root No Wt 1.0 h Col-0
## V44 Osmotic Stress 1.02 Root No Wt 3.0 h Col-0
## V46 Osmotic Stress 1.02 Root No Wt 6.0 h Col-0
## V48 Osmotic Stress 1.02 Root No Wt 12.0 h Col-0
## V50 Osmotic Stress 1.02 Root No Wt 24.0 h Col-0colnames(osmoroot) <- gsub(" Stress","",osmorootsampleinfo$timecourse)
# root12h
cols <- which(
sampleinfo$control =="No" &
sampleinfo$timecourse =="12.0 h" &
sampleinfo$tissue =="Root" )
root12h <- efp[,cols]
root12hsampleinfo <- sampleinfo[cols,]
root12hsampleinfo## category age tissue control mutant timecourse alias
## V24 Cold Stress 1.02 Root No Wt 12.0 h Col-0
## V48 Osmotic Stress 1.02 Root No Wt 12.0 h Col-0
## V72 Salt Stress 1.02 Root No Wt 12.0 h Col-0
## V100 Drought Stress 1.02 Root No Wt 12.0 h Col-0
## V124 Genotoxic Stress 1.02 Root No Wt 12.0 h Col-0
## V148 Oxidative Stress 1.02 Root No Wt 12.0 h Col-0
## V176 UV-B Stress 1.02 Root No Wt 12.0 h Col-0
## V204 Wounding Stress 1.02 Root No Wt 12.0 h Col-0
## V236 Heat Stress 1.02 Root No Wt 12.0 h Col-0colnames(root12h) <- gsub(" Stress","",root12hsampleinfo$category)
# root24h
cols <- which(
sampleinfo$control =="No" &
sampleinfo$timecourse =="24.0 h" &
sampleinfo$tissue =="Root" )
root24h <- efp[,cols]
root24hsampleinfo <- sampleinfo[cols,]
root24hsampleinfo## category age tissue control mutant timecourse alias
## V26 Cold Stress 1.02 Root No Wt 24.0 h Col-0
## V50 Osmotic Stress 1.02 Root No Wt 24.0 h Col-0
## V74 Salt Stress 1.02 Root No Wt 24.0 h Col-0
## V102 Drought Stress 1.02 Root No Wt 24.0 h Col-0
## V126 Genotoxic Stress 1.02 Root No Wt 24.0 h Col-0
## V150 Oxidative Stress 1.02 Root No Wt 24.0 h Col-0
## V178 UV-B Stress 1.02 Root No Wt 24.0 h Col-0
## V206 Wounding Stress 1.02 Root No Wt 24.0 h Col-0
## V238 Heat Stress 1.02 Root No Wt 24.0 h Col-0colnames(root24h) <- gsub(" Stress","",root24hsampleinfo$category)
# shoot12h
cols <- which(
sampleinfo$control =="No" &
sampleinfo$timecourse =="12.0 h" &
sampleinfo$tissue =="Shoot" )
shoot12h <- efp[,cols]
shoot12hsampleinfo <- sampleinfo[cols,]
shoot12hsampleinfo## category age tissue control mutant timecourse alias
## V23 Cold Stress 1.02 Shoot No Wt 12.0 h Col-0
## V47 Osmotic Stress 1.02 Shoot No Wt 12.0 h Col-0
## V71 Salt Stress 1.02 Shoot No Wt 12.0 h Col-0
## V99 Drought Stress 1.02 Shoot No Wt 12.0 h Col-0
## V123 Genotoxic Stress 1.02 Shoot No Wt 12.0 h Col-0
## V147 Oxidative Stress 1.02 Shoot No Wt 12.0 h Col-0
## V175 UV-B Stress 1.02 Shoot No Wt 12.0 h Col-0
## V203 Wounding Stress 1.02 Shoot No Wt 12.0 h Col-0
## V235 Heat Stress 1.02 Shoot No Wt 12.0 h Col-0colnames(shoot12h) <- gsub(" Stress","",shoot12hsampleinfo$category)
# root24h
cols <- which(
sampleinfo$control =="No" &
sampleinfo$timecourse =="24.0 h" &
sampleinfo$tissue =="Shoot" )
shoot24h <- efp[,cols]
shoot24hsampleinfo <- sampleinfo[cols,]
shoot24hsampleinfo## category age tissue control mutant timecourse alias
## V25 Cold Stress 1.02 Shoot No Wt 24.0 h Col-0
## V49 Osmotic Stress 1.02 Shoot No Wt 24.0 h Col-0
## V73 Salt Stress 1.02 Shoot No Wt 24.0 h Col-0
## V101 Drought Stress 1.02 Shoot No Wt 24.0 h Col-0
## V125 Genotoxic Stress 1.02 Shoot No Wt 24.0 h Col-0
## V149 Oxidative Stress 1.02 Shoot No Wt 24.0 h Col-0
## V177 UV-B Stress 1.02 Shoot No Wt 24.0 h Col-0
## V205 Wounding Stress 1.02 Shoot No Wt 24.0 h Col-0
## V237 Heat Stress 1.02 Shoot No Wt 24.0 h Col-0colnames(shoot24h) <- gsub(" Stress","",shoot24hsampleinfo$category)Heatmap
Scale off
#colour palette
colfunc <- colorRampPalette(c("blue", "white", "red"))
# shoot
heatmap.2( as.matrix(shoot), col=colfunc(25),scale="none",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Shoot")
# with proper names
shoot2 <- shoot
rownames(shoot2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(shoot2), col=colfunc(25),scale="none",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Shoot")
# root
heatmap.2( as.matrix(root), col=colfunc(25),scale="none",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Root")
# with proper names
root2 <- root
rownames(root2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="none",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Root")
# osmoroot
heatmap.2( as.matrix(osmoroot), col=colfunc(25),scale="none",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Root osmotic stress")
# with proper names
root2 <- osmoroot
rownames(root2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="none",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Root osmotic stress")
#root12h
heatmap.2( as.matrix(root12h), col=colfunc(25),scale="none",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Root 12 hrs")
root2 <- root12h
rownames(root2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="none",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Root 12 hrs")
#root24h
heatmap.2( as.matrix(root24h), col=colfunc(25),scale="none",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Root 24 hrs")
root2 <- root24h
rownames(root2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="none",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Root 24 hrs")
#shoot12h
heatmap.2( as.matrix(shoot12h), col=colfunc(25),scale="none",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Shoot 12 hrs")
shoot2 <- shoot12h
rownames(shoot2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(shoot2), col=colfunc(25),scale="none",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Shoot 12 hrs")
#root24h
heatmap.2( as.matrix(shoot24h), col=colfunc(25),scale="none",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Shoot 24 hrs")
shoot2 <- shoot24h
rownames(shoot2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="none",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Shoot 24 hrs")
Scale on
#colour palette
colfunc <- colorRampPalette(c("blue", "white", "red"))
# shoot
heatmap.2( as.matrix(shoot), col=colfunc(25),scale="row",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Shoot")
# with proper names
shoot2 <- shoot
rownames(shoot2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(shoot2), col=colfunc(25),scale="row",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Shoot")
# root
heatmap.2( as.matrix(root), col=colfunc(25),scale="row",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Root")
# with proper names
root2 <- root
rownames(root2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="row",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Root")
# osmoroot
heatmap.2( as.matrix(osmoroot), col=colfunc(25),scale="row",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Root osmotic stress")
# with proper names
root2 <- osmoroot
rownames(root2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="row",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Root osmotic stress")
#root12h
heatmap.2( as.matrix(root12h), col=colfunc(25),scale="row",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Root 12 hrs")
root2 <- root12h
rownames(root2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="row",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Root 12 hrs")
#root24h
heatmap.2( as.matrix(root24h), col=colfunc(25),scale="row",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Root 24 hrs")
root2 <- root24h
rownames(root2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="row",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Root 24 hrs")
#shoot12h
heatmap.2( as.matrix(shoot12h), col=colfunc(25),scale="row",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Shoot 12 hrs")
shoot2 <- shoot12h
rownames(shoot2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(shoot2), col=colfunc(25),scale="row",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Shoot 12 hrs")
#root24h
heatmap.2( as.matrix(shoot24h), col=colfunc(25),scale="row",
trace="none",margins = c(6,20), cexRow=.6, cexCol=.8, main="Shoot 24 hrs")
shoot2 <- shoot24h
rownames(shoot2) <- paste(rownames(genenames),genenames$NAME)
heatmap.2( as.matrix(root2), col=colfunc(25),scale="row",
trace="none",margins = c(6,25), cexRow=.6, cexCol=.8, main="Shoot 24 hrs")
Session information
So you know what version of R and packages was used.
sessionInfo()## R version 4.0.3 (2020-10-10)
## Platform: x86_64-pc-linux-gnu (64-bit)
## Running under: Ubuntu 18.04.5 LTS
##
## Matrix products: default
## BLAS: /usr/lib/x86_64-linux-gnu/blas/libblas.so.3.7.1
## LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.7.1
##
## 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] stats graphics grDevices utils datasets methods base
##
## other attached packages:
## [1] gplots_3.1.0
##
## loaded via a namespace (and not attached):
## [1] gtools_3.8.2 digest_0.6.27 bitops_1.0-6 magrittr_1.5
## [5] evaluate_0.14 KernSmooth_2.23-18 rlang_0.4.8 stringi_1.5.3
## [9] rmarkdown_2.5 tools_4.0.3 stringr_1.4.0 xfun_0.19
## [13] yaml_2.2.1 compiler_4.0.3 caTools_1.18.0 htmltools_0.5.0
## [17] knitr_1.30