Commit e44f2b1e authored by Venkat Malladi's avatar Venkat Malladi
Browse files

update repository.

parent ab30322c
......@@ -333,7 +333,7 @@ boxplot(2^pc_up_activated[,5:7], col=(c("yellow","yellow","yellow")), main="E2 u
axis(2, at=seq(0,20,5))
dev.off()
# The universe of down regulated genes by E2, separated by only looking at ones that have >2 fold change depression upon PARP-1 vs unaffected
# The universe of down regulated genes by E2, separated by only looking at ones that have >4 fold change depression upon PARP-1 vs unaffected
pc_affected_down_fc_sig <- pc_affected_fc[(pc_de==-1 | (pc_de==-1 & pc_de_p==-1)),]
pc_down_suppressed <- pc_affected_down_fc_sig[which(pc_affected_down_fc_sig$pc_et_e.table.logFC >= 2),]
pc_down_mod_suppressed <- pc_affected_down_fc_sig[which(pc_affected_down_fc_sig$pc_et_e.table.logFC >=0 & pc_affected_down_fc_sig$pc_et_e.table.logFC < 2),]
......@@ -807,7 +807,7 @@ axis(2, at=seq(0,25,5))
dev.off()
# The universe of down regulated genes by E2, separated by only looking at ones that have >2 fold change depression upon PARP-1 vs unaffected
# The universe of down regulated genes by E2, separated by only looking at ones that have >4 fold change depression upon PARP-1 vs unaffected
lnc_affected_down_fc_sig <- lnc_affected_fc[(lnc_de==-1 | (lnc_de==-1 & lnc_de_p==-1)),]
lnc_down_suppressed <- lnc_affected_down_fc_sig[which(lnc_affected_down_fc_sig$lnc_et_e.table.logFC >= 2),]
lnc_down_mod_suppressed <- lnc_affected_down_fc_sig[which(lnc_affected_down_fc_sig$lnc_et_e.table.logFC >=0 & lnc_affected_down_fc_sig$lnc_et_e.table.logFC < 2),]
......
# PARP-1 and Luc knockdowns with/without E2 on MCF-7 Cells
PARP-1 and Luciferace knockdowns with/without E2 in MCF-7 Cells
==========================================================
## Experiments:
1. GRO-seq
......@@ -38,6 +39,12 @@
* PARP-1
## Requirements:
- bedtools v2.25.0
- UCSC tools
- bedops v2.2.14
-
## Scripts:
1. parse_gencode.sh - Parses gencode file to make Protein coding universe and long-non coding universe
2. GRO-seq.Rmd - GRO-seq R markdown Analysis
......
#!/usr/bin/perl
open (FILE, "MinusStrand_matrix.txt") || die "can't: $!";
open (OUTPUT,">GROSeqTags_PlusStrand_minus_combined.cdt") || die "can't: $!";
%hash=();
while(<FILE>)
{
$line = $_;
chomp($line);
@array=split(/\t/,"$line");
$hash{$array[0]}=$line
}
close FILE;
open (FILE, "PlusStrand_matrix.txt") || die "can't: $!";
while(<FILE>)
{
$line1 = $_;
chomp($line1);
@a1=split(/\t/,"$line1");
if(exists $hash{$a1[0]})
{
@v=();
print OUTPUT "$line1\n";
print OUTPUT "$a1[0]";
$val=$hash{$a1[0]};
@v=split(/\t/,"$val");
for ($i=1;$i<=$#v;$i++)
{
if($v[$i] > 0)
{
print OUTPUT "\t-$v[$i]";
}
else
{
print OUTPUT "\t$v[$i]";
}
}
print OUTPUT "\n";
}
}
install.packages("ggplot2")
install.packages("stringr")
install.packages("reshape2")
library(ggplot2)
library(stringr)
library(reshape2)
# Boxplot FOXA1 E2 + Parp1 Lost,maintained,gained
expReads_foxa1 <- mean(c(NROW(l_e_foxa1),NROW(p_e_foxa1)))
l_mg_l_e_foxa1<- runMetaGene(features=start(lost), reads=l_e_foxa1, size=100,
, normCounts=expReads_foxa1/NROW(l_e_foxa1),up = 2000, sampling=FALSE,
mc.cores=getOption("mc.cores"))
l_mg_p_e_foxa1<- runMetaGene(features=start(lost), reads=p_e_foxa1, size=100,
, normCounts=expReads_foxa1/NROW(p_e_foxa1),up = 2000, sampling=FALSE,
mc.cores=getOption("mc.cores"))
lost_foxa1 <- c(as.data.frame(l_mg_l_e_foxa1$sense),as.data.frame(l_mg_p_e_foxa1$sense))
names(lost_foxa1) <- c("E", "P+E")
df <- melt(lost_foxa1)
df$V1 <- "Affected"
m_mg_l_e_foxa1<- runMetaGene(features=maintained, reads=l_e_foxa1, size=100,
, normCounts=expReads_foxa1/NROW(l_e_foxa1),up = 2000, sampling=FALSE,
mc.cores=getOption("mc.cores"))
m_mg_p_e_foxa1<- runMetaGene(features=maintained, reads=p_e_foxa1, size=100,
, normCounts=expReads_foxa1/NROW(p_e_foxa1),up = 2000, sampling=FALSE,
mc.cores=getOption("mc.cores"))
maintained_foxa1 <- c(as.data.frame(m_mg_l_e_foxa1$sense),as.data.frame(m_mg_p_e_foxa1$sense))
names(maintained_foxa1) <- c("E", "P+E")
df_2 <- melt(maintained_foxa1)
df_2$V1 <- "Affected"
g_mg_l_e_foxa1<- runMetaGene(features=gained, reads=l_e_foxa1, size=100,
, normCounts=expReads_foxa1/NROW(l_e_foxa1),up = 2000, sampling=FALSE,
mc.cores=getOption("mc.cores"))
g_mg_p_e_foxa1<- runMetaGene(features=gained, reads=p_e_foxa1, size=100,
, normCounts=expReads_foxa1/NROW(p_e_foxa1),up = 2000, sampling=FALSE,
mc.cores=getOption("mc.cores"))
gained_foxa1 <- c(as.data.frame(g_mg_l_e_foxa1$sense),as.data.frame(g_mg_p_e_foxa1$sense))
names(gained_foxa1) <- c("E", "P+E")
df_3 <- melt(gained_foxa1)
df_3$V1 <- "Unaffected"
df_4 <- rbind(df,df_2,df_3)
p <- ggplot(df_4, aes(x=V1, y=value, fill=L1)) + stat_boxplot(geom ='errorbar',linetype="F1",position=position_dodge(1)) + geom_boxplot(lwd=1,outlier.shape = NA,coef = 0, outlier.size = 0, position=position_dodge(1)) + scale_fill_manual(values=c("green4", "blue")) + labs(y="Norm. Read Counts",x="Status") + theme_bw() + theme(panel.grid.major = element_blank(),panel.grid.minor = element_blank(),axis.ticks=element_line(size=1), panel.border =element_rect(colour = "black",size=1)) + scale_y_continuous(limits=c(15,55))
jpeg('sgkolmen_foxa1_figure.jpg')
p
dev.off()
# Box plot
lost <- import.bed(con="lost_peaks-FOXA1.merge.bed")
maintained <- import.bed(con="maintained_peaks-FOXA1.merge.bed")
gained <- import.bed(con="gained_peaks-FOXA1.merge.bed")
bedtools multicov -bams ChIP-seq_Luc_KD_E2_FOXA1.bam ChIP-seq_PARP-1_KD_E2_FOXA1.bam -bed lost_peaks-FOXA1.merge.bed > lost_coverage-FOXA1
bedtools multicov -bams ChIP-seq_Luc_KD_E2_FOXA1.bam ChIP-seq_PARP-1_KD_E2_FOXA1.bam -bed maintained_peaks-FOXA1.merge.bed > maintained_coverage-FOXA1
bedtools multicov -bams ChIP-seq_Luc_KD_E2_FOXA1.bam ChIP-seq_PARP-1_KD_E2_FOXA1.bam -bed gained_peaks-FOXA1.merge.bed > gained_coverage-FOXA1
# RPKM
r_l_e_foxa1 <- NROW(l_e_foxa1)/1000000
r_p_e_foxa1 <- NROW(p_e_foxa1)/1000000
lost_counts <- read.table("/Volumes/work/GCRB/s163035/Par1-kd/m-g-l/lost_coverage-FOXA1", sep="\t")
lost_width <- width(lost)/1000
lost_foxa1 <- c(as.data.frame(lost_counts$V4/lost_width/r_l_e_foxa1),as.data.frame(lost_counts$V5/lost_width/r_l_e_foxa1))
names(lost_foxa1) <- c("E", "P+E")
maintained_counts <- read.table("/Volumes/work/GCRB/s163035/Par1-kd/m-g-l/maintained_coverage-FOXA1", sep="\t")
maintained_width <- width(maintained)/1000
maintained_foxa1 <- c(as.data.frame(maintained_counts$V4/maintained_width/r_l_e_foxa1),as.data.frame(maintained_counts$V5/maintained_width/r_l_e_foxa1))
names(maintained_foxa1) <- c("E", "P+E")
gained_counts <- read.table("/Volumes/work/GCRB/s163035/Par1-kd/m-g-l/gained_coverage-FOXA1", sep="\t")
gained_width <- width(gained)/1000
gained_foxa1 <- c(as.data.frame(gained_counts$V4/gained_width/r_l_e_foxa1),as.data.frame(gained_counts$V5/gained_width/r_l_e_foxa1))
names(gained_foxa1) <- c("E", "P+E")
df <- melt(lost_foxa1)
df$V1 <- "a-lost"
df_2 <- melt(maintained_foxa1)
df_2$V1 <- "b-maintained"
df_3 <- melt(gained_foxa1)
df_3$V1 <- "c-gained"
setwd("/Users/venkatmalladi/Projects/PARP-1_kd")
df_4 <- rbind(df,df_2,df_3)
p <- ggplot(df_4, aes(x=V1, y=value, fill=L1)) + stat_boxplot(geom ='errorbar',linetype="F1",position=position_dodge(1)) + geom_boxplot(lwd=1,outlier.shape = NA,coef = 0, outlier.size = 0, position=position_dodge(1)) + scale_fill_manual(values=c("green4", "blue")) + labs(y="RPKM",x="Status") + theme_bw() + theme(panel.grid.major = element_blank(),panel.grid.minor = element_blank(),axis.ticks=element_line(size=1), panel.border =element_rect(colour = "black",size=1)) + scale_y_continuous(limits=c(0,20))
jpeg('sgkolmen_foxa1_figure.jpg')
p
dev.off()
annotatePeaks.pl /Volumes/work/GCRB/s163035/Par1-kd/m-g-l/affected_peaks-FOXA1.merge.bed hg19 -size 4000 -hist 100 -ghist -d LUC-KD-Veh-FOXA1/ Luc-KD-E2-FOXA1/ PARP-1-KD-Veh-FOXA1/ PARP-1-KD-E2-FOXA1/ > foxa1_affected.txt
annotatePeaks.pl /Volumes/work/GCRB/s163035/Par1-kd/m-g-l/gained_peaks-FOXA1.merge.bed hg19 -size 4000 -hist 100 -ghist -d LUC-KD-Veh-FOXA1/ Luc-KD-E2-FOXA1/ PARP-1-KD-Veh-FOXA1/ PARP-1-KD-E2-FOXA1/ > foxa1_unaffected.txt
annotatePeaks.pl /Volumes/work/GCRB/s163035/Par1-kd/m-g-l/lost_peaks-FOXA1.merge.bed hg19 -size 4000 -hist 100 -ghist -d LUC-KD-Veh-FOXA1/ Luc-KD-E2-FOXA1/ PARP-1-KD-Veh-FOXA1/ PARP-1-KD-E2-FOXA1/ > foxa1_lost.txt
annotatePeaks.pl /Volumes/work/GCRB/s163035/Par1-kd/m-g-l/maintained_peaks-FOXA1.merge.bed hg19 -size 4000 -hist 100 -ghist -d LUC-KD-Veh-FOXA1/ Luc-KD-E2-FOXA1/ PARP-1-KD-Veh-FOXA1/ PARP-1-KD-E2-FOXA1/ > foxa1_maintained.txt
annotatePeaks.pl /Volumes/work/GCRB/s163035/Par1-kd/m-g-l/gained_peaks-FOXA1.merge.bed hg19 -size 4000 -hist 100 -ghist -d LUC-KD-Veh-FOXA1/ Luc-KD-E2-FOXA1/ PARP-1-KD-Veh-FOXA1/ PARP-1-KD-E2-FOXA1/ > foxa1_gained.txt
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