RNA-seq-Li

李老师RNA-seq PRC1.6-Gene和PRC1.6-Repeat数据分析

读取要处理的文件

library(ggplot2)
library (VennDiagram)
file.list<-c("PRC1.6-Gene/Rif1.KO.diff.csv",
             "PRC1.6-Gene/RNF2.KO.diff.csv",
             "PRC1.6-Repeat/Rif1.KO.rmsk.diff.csv",
             "PRC1.6-Repeat/RNF2.KO.rmsk.diff.csv")

火山图统计上下调基因数

for(file in file.list){
  DEG<-read.table(file,sep=",",header = T)
  #log2FoldChange_cutoff <- with(DEG,mean(abs( log2FoldChange)) + 2*sd(abs( log2FoldChange)) )
  log2FoldChange_cutoff<-1
  DEG$change = as.factor(ifelse(DEG$pvalue < 0.05 & abs(DEG$log2FoldChange) > log2FoldChange_cutoff,
                                ifelse(DEG$log2FoldChange > log2FoldChange_cutoff ,'UP','DOWN'),'NOT')
  )
  #
  this_tile <- paste0('Cutoff for log2FoldChange is ',round(log2FoldChange_cutoff,3),
                      'Pvalue is 0.05',
                      '\nThe number of up gene is ',nrow(DEG[DEG$change =='UP',]) ,
                      '\nThe number of down gene is ',nrow(DEG[DEG$change =='DOWN',])
  )
  #
  g = ggplot(data=DEG, 
             aes(x=log2FoldChange, y=-log10(pvalue), 
                 color=change)) +
    geom_point(alpha=0.4, size=1.75) +
    theme_set(theme_set(theme_bw(base_size=20)))+
    xlab("log2 fold change") + ylab("-log10 p-value") +
    ggtitle( this_tile ) + theme(plot.title = element_text(size=15,hjust = 0.5))+
    scale_colour_manual(values = c('blue','black','red')) ## corresponding to the levels(res$change)
  print(g)
  ggsave(g,filename = paste(file,'cutoff1','volcano.png',sep="."))
}

log2FC cutoff=1，pvalue=0.05时，Rif1上调基因数为759，下调基因数为410

Rif1.KO.diff.csv.cutoff1.volcano.png

log2FC cutoff=1，pvalue=0.05时，RNF2上调基因数为1461，下调基因数为1189

RNF2.KO.diff.csv.cutoff1.volcano.png

根据数据，计算log2FC cutoff值

log2FC cutoff=1.105，pvalue=0.05时，Rif1上调基因数为635，下调基因数为227

Rif1.KO.diff.csv.volcano.png

log2FC cutoff=2.034，pvalue=0.05时，Rif1上调基因数为612，下调基因数为144

RNF2.KO.diff.csv.volcano.png

repeat数据计算出来的log2FC cutoff值为3左右，得到的差异重复太少了，故直接取1

log2FC cutoff=1，pvalue=0.05时，Rif1上调Repeat数为254，下调基因数为171

Rif1.KO.rmsk.diff.csv.cutoff1.volcano.png

log2FC cutoff=1，pvalue=0.05时，Rif1上调Repeat数为155，下调基因数为31

RNF2.KO.rmsk.diff.csv.cutoff1.volcano.png

韦恩图

file.list<-c("PRC1.6-Gene/Rif1.KO.diff.csv",
             "PRC1.6-Gene/RNF2.KO.diff.csv")
             #"PRC1.6-Repeat/Rif1.KO.rmsk.diff.csv",
             #"PRC1.6-Repeat/RNF2.KO.rmsk.diff.csv",
             
up.list<-list()
down.list<-list()
for(file in file.list){
  DEG<-read.table(file,sep=",",header = T)
  log2FoldChange_cutoff <- with(DEG,mean(abs( log2FoldChange)) + 2*sd(abs( log2FoldChange)) )
  #log2FoldChange_cutoff<-1
  DEG$change = as.factor(ifelse(DEG$pvalue < 0.05 & abs(DEG$log2FoldChange) > log2FoldChange_cutoff,
                                ifelse(DEG$log2FoldChange > log2FoldChange_cutoff ,'UP','DOWN'),'NOT')
  )
  #
  up.list[[substr(basename(file), start = 1, stop = 4)]]<-DEG[DEG$change=='UP',"X"]
  down.list[[substr(basename(file), start = 1, stop = 4)]]<-DEG[DEG$change=='DOWN',"X"]
}
#
venn.diagram(up.list,
             "up.png",
             height = 450, 
             width = 450, 
             resolution =300, 
             imagetype="png", 
             col="white", 
             fill=c(colors()[616], colors()[38]), 
             alpha=c(0.6, 0.6), lwd=c(1, 1), cex=0.6,
             cat.dist=c(-0.07, -0.07), cat.pos=c(300, 60), cat.cex=0.7,
             main="Venn diagram of Rif1 and RNF2 up genes",main.cex=0.5)
venn.diagram(down.list,
             "down.png",
             height = 450, 
             width = 450, 
             resolution =300, 
             imagetype="png", 
             col="white", 
             fill=c(colors()[616], colors()[38]), 
             alpha=c(0.6, 0.6), lwd=c(1, 1), cex=0.6,
             cat.dist=c(-0.07, -0.07), cat.pos=c(300, 60), cat.cex=0.7,
             main="Venn diagram of Rif1 and RNF2 down genes",main.cex=0.5)

上调差异基因

up.png

下调差异基因

down.png

上调差异repeat

up.repeat.png

下调差异repeat

down.repeat.png