Pipeline for 55 Ind

QTL analysis pipeline

x = wc -l /project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_filtered/Filtered_APApeaks_merged_allchrom_noMP.bed
122488


seq 1 122488 > peak.num.txt

sort -k1,1 -k2,2n Filtered_APApeaks_merged_allchrom_noMP.bed > Filtered_APApeaks_merged_allchrom_noMP.sort.bed

paste /project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_filtered/Filtered_APApeaks_merged_allchrom_noMP.sort.bed peak.num.txt | column -s $'\t' -t > temp
awk '{print $1 "\t" $2 "\t" $3 "\t" $7  "\t"  $4 "\t"  $5 "\t" $6}' temp >   /project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_filtered/Filtered_APApeaks_merged_allchrom_noMP.sort.named.bed

#cut the chr  

sed 's/^chr//' /project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_filtered/Filtered_APApeaks_merged_allchrom_noMP.sort.named.bed > /project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_filtered/Filtered_APApeaks_merged_allchrom_noMP.sort.named.noCHR.bed

fout = file("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/file_id_mapping_total_Transcript_head.txt",'w')
infile= open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.fixed.fc", "r")
for line, i in enumerate(infile):
    if line == 0:
        i_list=i.split()
        files= i_list[10:-2]
        for each in files:
            full = each.split("/")[7]
            samp= full.split("-")[2:4]
            lim="_"
            samp_st=lim.join(samp)
            outLine= full[:-1] + "\t" + samp_st
            fout.write(outLine + "\n")
fout.close()

- create_fileid_geneLocAnno_nuclear.py  

fout = file("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/file_id_mapping_nuclear_Transcript_head.txt",'w')
infile= open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.fixed.fc", "r")
for line, i in enumerate(infile):
    if line == 0:
        i_list=i.split()
        files= i_list[10:-2]
        for each in files:
            full = each.split("/")[7]
            samp= full.split("-")[2:4]
            lim="_"
            samp_st=lim.join(samp)
            outLine= full[:-1] + "\t" + samp_st
            fout.write(outLine + "\n")
fout.close()

library(tidyverse)

totalPeakUs=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.fixed.pheno.fc", header = T, stringsAsFactors = F) %>% separate(chrom, sep = ":", into = c("chr", "start", "end", "id")) %>% separate(id, sep="_", into=c("gene", "strand", "peak"))

nuclearPeakUs=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.fixed.pheno.fc", header = T, stringsAsFactors = F) %>% separate(chrom, sep = ":", into = c("chr", "start", "end", "id")) %>% separate(id, sep="_", into=c("gene", "strand", "peak"))

ind=colnames(totalPeakUs)[7:dim(totalPeakUs)[2]]
totalPeakUs_CountNum=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.fixed.pheno.CountsOnlyNumeric.txt", col.names = ind)

nuclearPeakUs_CountNum=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.fixed.pheno.CountsOnlyNumeric.txt", col.names = ind)


#numeric with anno
totalPeak=as.data.frame(cbind(totalPeakUs[,1:6], totalPeakUs_CountNum))
nuclearPeak=as.data.frame(cbind(nuclearPeakUs[,1:6], nuclearPeakUs_CountNum))

#mean
totalPeakUs_CountNum_mean=rowMeans(totalPeakUs_CountNum)
nuclearPeakUs_CountNum_mean=rowMeans(nuclearPeakUs_CountNum)


#append mean to anno
TotalPeakUSMean=as.data.frame(cbind(totalPeakUs[,1:6],totalPeakUs_CountNum_mean))
NuclearPeakUSMean=as.data.frame(cbind(nuclearPeakUs[,1:6],nuclearPeakUs_CountNum_mean))

NuclearPeakUSMean_5perc=NuclearPeakUSMean %>% filter(nuclearPeakUs_CountNum_mean>=.05)
write.table(NuclearPeakUSMean_5perc,file="/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno.NoMP_sm_quant.Nuclear_fixed.pheno.5percPeaks.txt", row.names=F, col.names=F, quote = F)


TotalPeakUSMean_5per= TotalPeakUSMean %>% filter(totalPeakUs_CountNum_mean>=.05) 
write.table(TotalPeakUSMean_5per,file="/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno.NoMP_sm_quant.Total_fixed.pheno.5percPeaks.txt", row.names=F, col.names=F, quote = F)

#!/bin/bash

#SBATCH --job-name=run_filter_5percUsagePeaks
#SBATCH --account=pi-yangili1
#SBATCH --time=24:00:00
#SBATCH --output=run_filter_5percUsagePeaks.out
#SBATCH --error=run_filter_5percUsagePeaks.err
#SBATCH --partition=broadwl
#SBATCH --mem=12G
#SBATCH --mail-type=END

module load Anaconda3
source activate three-prime-env


Rscript filter_5percUsagePeaks.R

#zip file 
gzip filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.fixed.pheno_5perc.fc
gzip filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.fixed.pheno_5perc.fc


module load python
#leafcutter script
python /project2/gilad/briana/threeprimeseq/code/prepare_phenotype_table.py filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.fixed.pheno_5perc.fc.gz

python /project2/gilad/briana/threeprimeseq/code/prepare_phenotype_table.py filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.fixed.pheno_5perc.fc.gz




#source activate three-prime-env
 sh filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.fixed.pheno_5perc.fc.gz_prepare.sh
sh filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.fixed.pheno_5perc.fc.gz_prepare.sh


#keep only 2 PCs
head -n 3 filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.fixed.pheno_5perc.fc.gz.PCs > filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.fixed.pheno_5perc.fc.gz.2PCs
head -n 3 filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.fixed.pheno_5perc.fc.gz.PCs > filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.fixed.pheno_5perc.fc.gz.2PCs

#make a sample list  

fout = open("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs/SAMPLE.txt",'w')

for ln in open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/file_id_mapping_total_Transcript_head.txt", "r"):
    bam, sample = ln.split()
    line=sample[:-2]
    fout.write("NA"+line + "\n")
fout.close()

totQTLs=read.table("../data/perm_QTL_GeneLocAnno_noMP_5percov/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno.NoMP_sm_quant.Total.fixed.pheno_5perc_permResBH.txt", stringsAsFactors = F, header=T)

Sig_TotQTLs= totQTLs %>% filter(-log10(bh)>=1)
nrow(Sig_TotQTLs)

[1] 363

totQTLs %>% separate(pid, into=c("chr", "start", "end", "id"), sep=":") %>% separate(id, into=c("gene", "strand", "peak"), sep="_") %>% group_by(gene) %>% select(gene) %>% tally() %>% nrow()

Warning: Expected 3 pieces. Additional pieces discarded in 3 rows [815,
816, 817].

[1] 10633

nucQTLs=read.table("../data/perm_QTL_GeneLocAnno_noMP_5percov/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno.NoMP_sm_quant.Nuclear.fixed.pheno_5perc_permResBH.txt", stringsAsFactors = F, header=T)

Sig_NucQTLs= nucQTLs %>% filter(-log10(bh)>=1)
nrow(Sig_NucQTLs)

[1] 623

nucQTLs %>% separate(pid, into=c("chr", "start", "end", "id"), sep=":") %>% separate(id, into=c("gene", "strand", "peak"), sep="_") %>% group_by(gene) %>% select(gene) %>% tally() %>% nrow()

Warning: Expected 3 pieces. Additional pieces discarded in 3 rows [990,
991, 992].

[1] 10852

QTLs only peaks in the 3’ UTR

I want to subset to only look at the peaks that assign to the 3’ UTRs.

Remake the processGenLocPeakAnno2SAF.py code to only include peaks maping to a 3’ UTR

processGenLocPeakAnno2SAF_3UTRonly.py

inFile="/project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_GeneLoc/Filtered_APApeaks_merged_allchrom_noMP.sort.named.noCHR_geneLoc.bed"
outFile=open("/project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_GeneLoc_3UTR/Filtered_APApeaks_merged_allchrom_noMP.sort.named.noCHR_geneLocParsed_3UTR.SAF" , "w")

outFile.write("GeneID\tChr\tStart\tEnd\tStrand\n")
for ln in open(inFile, "r"):
    chrom, start, end, peak, cov, strand, score, anno = ln.split()
    if anno==".": 
        continue  
    anno_lst=anno.split(",")
    if len(anno_lst)==1:
        gene=anno_lst[0].split(":")[1]
        if anno_lst[0].split(":")[0]=="utr3":
            peak_i=int(peak)
            start_i=int(start)
            end_i=int(end)
            ID="peak%d:%s:%d:%d:%s:%s"%(peak_i, chrom, start_i, end_i, strand, gene)
            outFile.write("%s\t%s\t%d\t%d\t%s\n"%(ID, chrom, start_i, end_i, strand))
    else:
        type_dic={}
        for each in anno_lst:
            type_dic[each.split(":")[0]]=each.split(":")[1]
        if "utr3" in type_dic.keys():
            gene=type_dic["utr3"]
            peak_i=int(peak)
            start_i=int(start)
            end_i=int(end)
            ID="peak%d:%s:%d:%d:%s:%s"%(peak_i, chrom, start_i, end_i, strand, gene)
            outFile.write("%s\t%s\t%d\t%d\t%s\n"%(ID, chrom, start_i, end_i, strand))
outFile.close()

This results in 28373 peaks.

• feature counts GeneLocAnno_fc_TN_noMP_3UTR.sh

#!/bin/bash

#SBATCH --job-name=GeneLocAnno_fc_TN_noMP_3UTR
#SBATCH --account=pi-yangili1
#SBATCH --time=24:00:00
#SBATCH --output=GeneLocAnno_fc_TN_noMP_3UTR.out
#SBATCH --error=GeneLocAnno_fc_TN_noMP_3UTR.err
#SBATCH --partition=broadwl
#SBATCH --mem=12G
#SBATCH --mail-type=END

module load Anaconda3
source activate three-prime-env

featureCounts -O -a /project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_GeneLoc_3UTR/Filtered_APApeaks_merged_allchrom_noMP.sort.named.noCHR_geneLocParsed_3UTR.SAF -F SAF -o /project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fc /project2/gilad/briana/threeprimeseq/data/bam_NoMP_sort/*T-combined-sort.noMP.sort.bam -s 1

featureCounts -O -a /project2/gilad/briana/threeprimeseq/data/mergedPeaks_noMP_GeneLoc_3UTR/Filtered_APApeaks_merged_allchrom_noMP.sort.named.noCHR_geneLocParsed_3UTR.SAF -F SAF -o /project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fc /project2/gilad/briana/threeprimeseq/data/bam_NoMP_sort/*N-combined-sort.noMP.sort.bam -s 1

fix_head_fc_geneLoc_tot_noMP_3UTR.py

infile= open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fc", "r")
fout = open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.fc",'w')
for line, i in enumerate(infile):
    if line == 1:
        i_list=i.split()
        libraries=i_list[:6]
        for sample in i_list[6:]:
            full = sample.split("/")[7]
            samp= full.split("-")[2:4]
            lim="_"
            samp_st=lim.join(samp)
            libraries.append(samp_st)
        first_line= "\t".join(libraries)
        fout.write(first_line + '\n')
    else :
        fout.write(i)
fout.close()

fix header fix_head_fc_geneLoc_nuc_noMP_3UTR.py

infile= open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fc", "r")
fout = open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.fc",'w')
for line, i in enumerate(infile):
    if line == 1:
        i_list=i.split()
        libraries=i_list[:6]
        for sample in i_list[6:]:
            full = sample.split("/")[7]
            samp= full.split("-")[2:4]
            lim="_"
            samp_st=lim.join(samp)
            libraries.append(samp_st)
        first_line= "\t".join(libraries)
        fout.write(first_line + '\n')
    else :
        fout.write(i)
fout.close()

File IDs:
* /project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/file_id_mapping_nuclear_Transcript_head.txt * c/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/file_id_mapping_total_Transcript_head.txt

• make phenotype run_makePhen_sep_GeneLocAnno_noMP_3UTR.sh

These are also different. The start and end for calling QTLs will be the peak rather than the gene start.

makePhenoRefSeqPeaks_GeneLoc_Total_noMP_3UTR.py

#PYTHON 3

dic_IND = {}
dic_BAM = {}

for ln in open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/file_id_mapping_total_Transcript_head.txt"):
    bam, IND = ln.split("\t")
    IND = IND.strip()
    dic_IND[bam] = IND
    if IND not in dic_BAM:
        dic_BAM[IND] = []
    dic_BAM[IND].append(bam)


#now I have ind dic with keys as the bam and ind as the values
#I also have a bam dic with ind as the keys and bam as the values  
    
inds=list(dic_BAM.keys()) #list of ind libraries  

#gene start and end dictionaries: 
dic_geneS = {}
dic_geneE = {}
for ln in open("/project2/gilad/briana/genome_anotation_data/RefSeq_annotations/ncbiRefSeq_endAllGenes.sort.bed"):
    chrom, start, end, geneID, score, strand = ln.split('\t')
    gene= geneID.split(":")[1]
#    if "-" in gene:
 #       gene=gene.split("-")[0]
    if gene not in dic_geneS:
        dic_geneS[gene]=int(start)
        dic_geneE[gene]=int(end)
        


#list of genes   

count_file=open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.fc", "r")
genes=[]
for line , i in enumerate(count_file):
    if line > 1:
        i_list=i.split()
        id=i_list[0]
        id_list=id.split(":")
        gene=id_list[5]
        if gene not in genes:
            genes.append(gene)
            
#make the ind and gene dic  
dic_dub={}
for g in genes:
    dic_dub[g]={}
    for i in inds:
        dic_dub[g][i]=0


#populate the dictionary  
count_file=open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.fc", "r")
for line, i in enumerate(count_file):
    if line > 1:
        i_list=i.split()
        id=i_list[0]
        id_list=id.split(":")
        g= id_list[5]
        values=list(i_list[6:])
        list_list=[]
        for ind,val in zip(inds, values):
            list_list.append([ind, val])
        for num, name in enumerate(list_list):
            dic_dub[g][list_list[num][0]] += int(list_list[num][1])
        

#write the file by acessing the dictionary and putting values in the table ver the value in the dic 
        

fout=open("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno.fc","w")
peak=["chrom"]
inds_noL=[]
for each in inds:
    indsNA= "NA" + each[:-2]
    inds_noL.append(indsNA) 
fout.write(" ".join(peak + inds_noL) + '\n' )


count_file=open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.fc", "r")
for line , i in enumerate(count_file):
    if line > 1:
        i_list=i.split()
        id=i_list[0]
        id_list=id.split(":")
        gene=int(id_list[5])
        #start=dic_geneS[id_list[5]]
        start=int(id_list[2])
        #end=dic_geneE[id_list[5]]
        end=id_list[3]
        buff=[]
        buff.append("chr%s:%d:%d:%s_%s_%s"%(id_list[1], start, end, id_list[5], id_list[4], id_list[0]))
        for x,y in zip(i_list[6:], inds):
            b=int(dic_dub[gene][y])
            t=int(x)
            buff.append("%d/%d"%(t,b))
        fout.write(" ".join(buff)+ '\n')
        
fout.close()

makePhenoRefSeqPeaks_GeneLoc_Nuclear_noMP_3UTR.py

#PYTHON 3

dic_IND = {}
dic_BAM = {}

for ln in open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno/file_id_mapping_nuclear_Transcript_head.txt"):
    bam, IND = ln.split("\t")
    IND = IND.strip()
    dic_IND[bam] = IND
    if IND not in dic_BAM:
        dic_BAM[IND] = []
    dic_BAM[IND].append(bam)


#now I have ind dic with keys as the bam and ind as the values
#I also have a bam dic with ind as the keys and bam as the values  
    
inds=list(dic_BAM.keys()) #list of ind libraries  

#gene start and end dictionaries: 
dic_geneS = {}
dic_geneE = {}
for ln in open("/project2/gilad/briana/genome_anotation_data/RefSeq_annotations/ncbiRefSeq_endAllGenes.sort.bed"):
    chrom, start, end, geneID, score, strand = ln.split('\t')
    gene= geneID.split(":")[1]
#    if "-" in gene:
 #       gene=gene.split("-")[0]
    if gene not in dic_geneS:
        dic_geneS[gene]=int(start)
        dic_geneE[gene]=int(end)
        


#list of genes   

count_file=open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.fc", "r")
genes=[]
for line , i in enumerate(count_file):
    if line > 1:
        i_list=i.split()
        id=i_list[0]
        id_list=id.split(":")
        gene=id_list[5]
        if gene not in genes:
            genes.append(gene)
            
#make the ind and gene dic  
dic_dub={}
for g in genes:
    dic_dub[g]={}
    for i in inds:
        dic_dub[g][i]=0


#populate the dictionary  
count_file=open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.fc", "r")
for line, i in enumerate(count_file):
    if line > 1:
        i_list=i.split()
        id=i_list[0]
        id_list=id.split(":")
        g= id_list[5]
        values=list(i_list[6:])
        list_list=[]
        for ind,val in zip(inds, values):
            list_list.append([ind, val])
        for num, name in enumerate(list_list):
            dic_dub[g][list_list[num][0]] += int(list_list[num][1])
        

#write the file by acessing the dictionary and putting values in the table ver the value in the dic 
        

fout=open("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno.fc","w")
peak=["chrom"]
inds_noL=[]
for each in inds:
    indsNA= "NA" + each[:-2]
    inds_noL.append(indsNA) 
fout.write(" ".join(peak + inds_noL) + '\n' )


count_file=open("/project2/gilad/briana/threeprimeseq/data/filtPeakOppstrand_cov_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.fc", "r")
for line , i in enumerate(count_file):
    if line > 1:
        i_list=i.split()
        id=i_list[0]
        id_list=id.split(":")
        gene=id_list[5]
        #start=dic_geneS[id_list[5]]
        start=int(id_list[2])
        #end=dic_geneE[id_list[5]]
        end=int(id_list[3])
        buff=[]
        buff.append("chr%s:%d:%d:%s_%s_%s"%(id_list[1], start, end, id_list[5], id_list[4], id_list[0]))
        for x,y in zip(i_list[6:], inds):
            b=int(dic_dub[gene][y])
            t=int(x)
            buff.append("%d/%d"%(t,b))
        fout.write(" ".join(buff)+ '\n')
        
fout.close()

• convert to usage pheno2CountOnly_genelocAnno_3UTR.R

library(reshape2)
library(tidyverse)


totalPeakUs=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno.fc", header = T, stringsAsFactors = F) %>% separate(chrom, sep = ":", into = c("chr", "start", "end", "id")) %>% separate(id, sep="_", into=c("gene", "strand", "peak"))
nuclearPeakUs=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno.fc", header = T, stringsAsFactors = F) %>% separate(chrom, sep = ":", into = c("chr", "start", "end", "id")) %>% separate(id, sep="_", into=c("gene", "strand", "peak"))



write.table(totalPeakUs[,7:dim(totalPeakUs)[2]], file="/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno.CountsOnly",quote=FALSE, col.names = F, row.names = F)

write.table(nuclearPeakUs[,7:dim(nuclearPeakUs)[2]], file="/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno.CountsOnly",quote=FALSE, col.names = F, row.names = F)

- counts to numeric convertCount2Numeric_noMP_GeneLocAnno_3UTR.py   

def convert(infile, outfile):
  final=open(outfile, "w")
  for ln in open(infile, "r"):
    line_list=ln.split()
    new_list=[]
    for i in line_list:
      num, dem = i.split("/")
      if dem == "0":
        perc = "0.00"
      else:
        perc = int(num)/int(dem)
        perc=round(perc,2)
        perc= str(perc)
      new_list.append(perc)
    final.write("\t".join(new_list)+ '\n')
  final.close()
  
convert("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno.CountsOnly","/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno.CountsOnlyNumeric.txt")


convert("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno.CountsOnly","/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno.CountsOnlyNumeric.txt")

filter_5percUsagePeaks_3UTR.R

library(tidyverse)

totalPeakUs=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno.fc", header = T, stringsAsFactors = F) %>% separate(chrom, sep = ":", into = c("chr", "start", "end", "id")) %>% separate(id, sep="_", into=c("gene", "strand", "peak"))

nuclearPeakUs=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno.fc", header = T, stringsAsFactors = F) %>% separate(chrom, sep = ":", into = c("chr", "start", "end", "id")) %>% separate(id, sep="_", into=c("gene", "strand", "peak"))

ind=colnames(totalPeakUs)[7:dim(totalPeakUs)[2]]
totalPeakUs_CountNum=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno.CountsOnlyNumeric.txt", col.names = ind)

nuclearPeakUs_CountNum=read.table("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno.CountsOnlyNumeric.txt", col.names = ind)


#numeric with anno
totalPeak=as.data.frame(cbind(totalPeakUs[,1:6], totalPeakUs_CountNum))
nuclearPeak=as.data.frame(cbind(nuclearPeakUs[,1:6], nuclearPeakUs_CountNum))

#mean
totalPeakUs_CountNum_mean=rowMeans(totalPeakUs_CountNum)
nuclearPeakUs_CountNum_mean=rowMeans(nuclearPeakUs_CountNum)


#append mean to anno
TotalPeakUSMean=as.data.frame(cbind(totalPeakUs[,1:6],totalPeakUs_CountNum_mean))
NuclearPeakUSMean=as.data.frame(cbind(nuclearPeakUs[,1:6],nuclearPeakUs_CountNum_mean))

NuclearPeakUSMean_5perc=NuclearPeakUSMean %>% filter(nuclearPeakUs_CountNum_mean>=.05)
write.table(NuclearPeakUSMean_5perc,file="/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno.NoMP_sm_quant.Nuclear.3UTR_fixed.pheno.5percPeaks.txt", row.names=F, col.names=F, quote = F)


TotalPeakUSMean_5per= TotalPeakUSMean %>% filter(totalPeakUs_CountNum_mean>=.05) 
write.table(TotalPeakUSMean_5per,file="/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno.NoMP_sm_quant.Total.3UTR_fixed.pheno.5percPeaks.txt", row.names=F, col.names=F, quote = F)

• run_filter_5percUsagePeaks_3UTR.sh

#!/bin/bash

#SBATCH --job-name=run_filter_5percUsagePeaks_3UTR
#SBATCH --account=pi-yangili1
#SBATCH --time=24:00:00
#SBATCH --output=run_filter_5percUsagePeaks_3UTR.out
#SBATCH --error=run_filter_5percUsagePeaks_3UTR.err
#SBATCH --partition=broadwl
#SBATCH --mem=12G
#SBATCH --mail-type=END

module load Anaconda3
source activate three-prime-env


Rscript filter_5percUsagePeaks_3UTR.R

filterPheno_bothFraction_GeneLocAnno_5perc_3UTR.py

#python  

totalokPeaks5perc_file="/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno.NoMP_sm_quant.Total.3UTR_fixed.pheno.5percPeaks.txt"

totalokPeaks5perc={}
for ln in open(totalokPeaks5perc_file,"r"):
    peakname=ln.split()[5]
    totalokPeaks5perc[peakname]=""


nuclearokPeaks5perc_file="/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno.NoMP_sm_quant.Nuclear.3UTR_fixed.pheno.5percPeaks.txt"
nuclearokPeaks5perc={}
for ln in open(nuclearokPeaks5perc_file,"r"):
    peakname=ln.split()[5]
    nuclearokPeaks5perc[peakname]=""
    
    
totalPhenoBefore=open("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno.fc","r")
totalPhenoAfter=open("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc", "w")
for num, ln in enumerate(totalPhenoBefore):
    if num ==0:
        totalPhenoAfter.write(ln)
    else:  
        id=ln.split()[0].split(":")[3].split("_")[2]
        if id in totalokPeaks5perc.keys():
            totalPhenoAfter.write(ln)
totalPhenoAfter.close()  

nuclearPhenoBefore=open("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno.fc","r")
nuclearPhenoAfter=open("/project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc", "w")
for num, ln in enumerate(nuclearPhenoBefore):
    if num ==0:
        nuclearPhenoAfter.write(ln)
    else:  
        id=ln.split()[0].split(":")[3].split("_")[2]
        if id in nuclearokPeaks5perc.keys():
            nuclearPhenoAfter.write(ln)
nuclearPhenoAfter.close() 

In /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/

#zip file 
gzip filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc
gzip filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc


module load python
#leafcutter script
python /project2/gilad/briana/threeprimeseq/code/prepare_phenotype_table.py filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz

python /project2/gilad/briana/threeprimeseq/code/prepare_phenotype_table.py filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz




#source activate three-prime-env
sh filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz_prepare.sh
sh filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz_prepare.sh


#keep only 2 PCs
head -n 3 filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz.PCs > filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz.2PCs
head -n 3 filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz.PCs > filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz.2PCs

QTL scripts:
Same smaple list

• APAqtl_nominal_GeneLocAnno_noMP_5percUsage_3UTR.sh

#!/bin/bash


#SBATCH --job-name=APAqtl_nominal_GeneLocAnno_noMP_5percUsage
#SBATCH --account=pi-yangili1
#SBATCH --time=24:00:00
#SBATCH --output=APAqtl_nominal_GeneLocAnno_noMP_5percUsage.out
#SBATCH --error=APAqtl_nominal_GeneLocAnno_noMP_5percUsage.err
#SBATCH --partition=broadwl
#SBATCH --mem=12G
#SBATCH --mail-type=END

for i in 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 
do
/home/brimittleman/software/bin/FastQTL/bin/fastQTL.static --vcf /project2/gilad/briana/YRI_geno_hg19/chr$i.dose.filt.vcf.gz --cov /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz.2PCs --bed /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz.qqnorm_chr$i.gz --out /project2/gilad/briana/threeprimeseq/data/nominal_APAqtl_GeneLocAnno_noMP_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz.qqnorm_chr$i.nominal.out --chunk 1 1  --window 5e5 --include-samples /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs/SAMPLE.txt
done


for i in 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 
do
/home/brimittleman/software/bin/FastQTL/bin/fastQTL.static --vcf /project2/gilad/briana/YRI_geno_hg19/chr$i.dose.filt.vcf.gz --cov /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz.2PCs --bed /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz.qqnorm_chr$i.gz --out /project2/gilad/briana/threeprimeseq/data/nominal_APAqtl_GeneLocAnno_noMP_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz.qqnorm_chr$i.nominal.out --chunk 1 1  --window 5e5 --include-samples /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs/SAMPLE.txt
done

• APAqtl_perm_GeneLocAnno_noMP_5percUsage_3UTR.sh

#!/bin/bash


#SBATCH --job-name=APAqtl_perm_GeneLocAnno_noMP_5percUsage
#SBATCH --account=pi-yangili1
#SBATCH --time=24:00:00
#SBATCH --output=APAqtl_perm_GeneLocAnno_noMP_5percUsage.out
#SBATCH --error=APAqtl_perm_GeneLocAnno_noMP_5percUsage.err
#SBATCH --partition=broadwl
#SBATCH --mem=12G
#SBATCH --mail-type=END

for i in 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 
do
/home/brimittleman/software/bin/FastQTL/bin/fastQTL.static  --permute 1000 --vcf /project2/gilad/briana/YRI_geno_hg19/chr$i.dose.filt.vcf.gz --cov /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz.2PCs --bed /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz.qqnorm_chr$i.gz --out /project2/gilad/briana/threeprimeseq/data/perm_APAqtl_GeneLocAnno_noMP_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc.fc.gz.qqnorm_chr$i.perm.out --chunk 1 1  --window 5e5 --include-samples /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs/SAMPLE.txt
done


for i in 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 
do
/home/brimittleman/software/bin/FastQTL/bin/fastQTL.static   --permute 1000 --vcf /project2/gilad/briana/YRI_geno_hg19/chr$i.dose.filt.vcf.gz --cov /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz.2PCs --bed /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz.qqnorm_chr$i.gz --out /project2/gilad/briana/threeprimeseq/data/perm_APAqtl_GeneLocAnno_noMP_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc.fc.gz.qqnorm_chr$i.perm.out --chunk 1 1  --window 5e5 --include-samples /project2/gilad/briana/threeprimeseq/data/phenotypes_filtPeakTranscript_noMP_GeneLocAnno_5percUs/SAMPLE.txt
done

• run_APAqtlpermCorrectQQplot_GeneLocAnno_noMP_5perUs_3UTR.sh

APAqtlpermCorrectQQplot_GeneLocAnno_noMP_5perUs_3UTR.R

library(dplyr)


##total results
tot.perm= read.table("/project2/gilad/briana/threeprimeseq/data/perm_APAqtl_GeneLocAnno_noMP_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc_permRes.txt",head=F, stringsAsFactors=F, col.names = c("pid", "nvar", "shape1", "shape2", "dummy", "sid", "dist", "npval", "slope", "ppval", "bpval"))

#BH correction
tot.perm$bh=p.adjust(tot.perm$bpval, method="fdr")

#plot qqplot
png("/project2/gilad/briana/threeprimeseq/output/plots/qqplot_total_APAperm_GeneLocAnno_noMP_5percCov_3UTR.png") 
qqplot_total= qqplot(-log10(runif(nrow(tot.perm))), -log10(tot.perm$bpval),ylab="-log10 Total permuted pvalue", xlab="Uniform expectation", main="Total permuted pvalues for all snps\n Gene Loc Anno")
abline(0,1)
dev.off()

#write df with BH  

write.table(tot.perm, file = "/project2/gilad/briana/threeprimeseq/data/perm_APAqtl_GeneLocAnno_noMP_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc_permResBH.txt", col.names = T, row.names = F, quote = F)

##nuclear results  


nuc.perm= read.table("/project2/gilad/briana/threeprimeseq/data/perm_APAqtl_GeneLocAnno_noMP_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc_permRes.txt",head=F, stringsAsFactors=F, col.names = c("pid", "nvar", "shape1", "shape2", "dummy", "sid", "dist", "npval", "slope", "ppval", "bpval"))
nuc.perm$bh=p.adjust(nuc.perm$bpval, method="fdr")


#plot qqplot
png("/project2/gilad/briana/threeprimeseq/output/plots/qqplot_nuclear_APAperm_GeneLocAnno_noMP_5percCov_3UTR.png") 
qqplot(-log10(runif(nrow(nuc.perm))), -log10(nuc.perm$bpval),ylab="-log10 Nuclear permuted pvalue", xlab="Uniform expectation", main="Nuclear permuted pvalues for all snps \n Gene Loc Anno")
abline(0,1)
dev.off()

# write df with BH
write.table(nuc.perm, file = "/project2/gilad/briana/threeprimeseq/data/perm_APAqtl_GeneLocAnno_noMP_5percUs_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc_permResBH.txt", col.names = T, row.names = F, quote = F)

totQTLs_UTR=read.table("../data/perm_QTL_GeneLocAnno_noMP_5percov_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Total.3UTR.fixed.pheno_5perc_permResBH.txt", stringsAsFactors = F, header=T)

Sig_TotQTLs_UTR= totQTLs_UTR %>% filter(-log10(bh)>=1)
nrow(Sig_TotQTLs_UTR)

[1] 114

How many genes are tested:

totQTLs_UTR %>% separate(pid, into=c("chr", "start", "end", "id"), sep=":") %>% separate(id, into=c("gene", "strand", "peak"), sep="_") %>% group_by(gene) %>% select(gene) %>% tally() %>% nrow()

[1] 6313

nucQTLs_UTR=read.table("../data/perm_QTL_GeneLocAnno_noMP_5percov_3UTR/filtered_APApeaks_merged_allchrom_refseqGenes.GeneLocAnno_NoMP_sm_quant.Nuclear.3UTR.fixed.pheno_5perc_permResBH.txt", stringsAsFactors = F, header=T)

Sig_NucQTLs_UTR= nucQTLs_UTR %>% filter(-log10(bh)>=1)
nrow(Sig_NucQTLs_UTR)

[1] 108

How many genes:

nucQTLs_UTR %>% separate(pid, into=c("chr", "start", "end", "id"), sep=":") %>% separate(id, into=c("gene", "strand", "peak"), sep="_") %>% group_by(gene) %>% select(gene) %>% tally() %>% nrow()

[1] 6349

Diff Isoform Pipeline:

Pipeline comes from New gene Annodation Dif Iso
* filternamePeaks5percCov_GeneLocAnno.py * bothFrac_processed_GeneLocAnno_FC.sh * fix_head_fc_procBothFrac_GeneLocAnno.py * fc2leafphen_processed_GeneLocAnno.py
* subset_diffisopheno_processed_GeneLocAnno.py/ run_subset_diffisopheno_processed_GeneLocAnno.sh * makeLCSampleList_processed_GeneLocAnno.py
* run_leafcutter_ds_bychrom_processed_GeneLocAnno.sh

awk '{if(NR>1)print}' /project2/gilad/briana/threeprimeseq/data/diff_iso_processed_GeneLocAnno/TN_diff_isoform_GeneLocAnno_chr*.txt_effect_sizes.txt > /project2/gilad/briana/threeprimeseq/data/diff_iso_processed_GeneLocAnno/TN_diff_isoform_GeneLocAnno_AllChrom.txt_effect_sizes.txt

awk '{if(NR>1)print}' /project2/gilad/briana/threeprimeseq/data/diff_iso_processed_GeneLocAnno/TN_diff_isoform_GeneLocAnno_chr*cluster_significance.txt > /project2/gilad/briana/threeprimeseq/data/diff_iso_processed_GeneLocAnno/TN_diff_isoform_GeneLocAnno_AllChrom.txt_cluster_significance.txt

diffIso=read.table("../data/diff_iso_GeneLocAnno/TN_diff_isoform_GeneLocAnno_AllChrom.txt_cluster_significance.txt", header = F,col.names = c("status",   "loglr",    "df",   "p",    "cluster",  "p.adjust"),stringsAsFactors = F,sep="\t") %>% filter(status == "Success")


diffIso$p.adjust=as.numeric(as.character(diffIso$p.adjust))


qqplot(-log10(runif(nrow(diffIso))), -log10(diffIso$p.adjust),ylab="-log10 Total Adjusted Leafcutter pvalue", xlab="-log 10 Uniform expectation", main="Leafcutter differencial isoform analysis between fractions")
abline(0,1)

effectsize=read.table("../data/diff_iso_GeneLocAnno/TN_diff_isoform_GeneLocAnno_AllChrom.txt_effect_sizes.txt", stringsAsFactors = F, col.names=c('intron',  'logef' ,'Nuclear', 'Total','deltapsi'))

effectsize$deltapsi=as.numeric(as.character(effectsize$deltapsi))

Warning: NAs introduced by coercion

filterHighPSI=effectsize %>% filter(abs(deltapsi)>.4) %>% arrange(deltapsi)
slice(filterHighPSI, 1:10)

                              intron              logef           Nuclear
1       chr16:81123538:81123627:GCSH  -1.79087571426241 0.724883044705174
2     chr3:25705115:25705187:MIR4442  -1.32432047353345 0.660590394608791
3         chr6:84007319:84007404:ME1  -1.68685519425574 0.671530253986571
4  chr19:36806427:36806515:LINC00665  -1.17617088053315 0.809837765186814
5      chr14:47718556:47718593:MDGA2   -1.1825787812967 0.659822642518761
6      chr14:51976014:51976057:FRMD6  -1.56759469994778 0.636611882071359
7  chr20:58647920:58648008:C20orf197  -1.09984402063945 0.691727788494324
8      chr6:37812035:37812094:ZFAND3  -1.61085385492872 0.617570611488878
9     chr8:104953022:104953097:RIMS2  -1.39009184488063 0.611545558430189
10  chr7:147596894:147596932:CNTNAP2 -0.827276301229926  0.72731101101958
               Total   deltapsi
1  0.151199900846243 -0.5736831
2  0.121029754316441 -0.5395606
3  0.134625920458626 -0.5369043
4   0.28835529406143 -0.5214825
5  0.154119006269487 -0.5057036
6  0.139375449680577 -0.4972364
7  0.199171791192942 -0.4925560
8  0.127058493301525 -0.4905121
9   0.13467815341195 -0.4768674
10 0.259840841892183 -0.4674702

Session information

sessionInfo()

R version 3.5.1 (2018-07-02)
Platform: x86_64-apple-darwin15.6.0 (64-bit)
Running under: macOS  10.14.1

Matrix products: default
BLAS: /Library/Frameworks/R.framework/Versions/3.5/Resources/lib/libRblas.0.dylib
LAPACK: /Library/Frameworks/R.framework/Versions/3.5/Resources/lib/libRlapack.dylib

locale:
[1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     

other attached packages:
 [1] bindrcpp_0.2.2  workflowr_1.1.1 forcats_0.3.0   stringr_1.3.1  
 [5] dplyr_0.7.6     purrr_0.2.5     readr_1.1.1     tidyr_0.8.1    
 [9] tibble_1.4.2    ggplot2_3.0.0   tidyverse_1.2.1

loaded via a namespace (and not attached):
 [1] Rcpp_0.12.19      cellranger_1.1.0  plyr_1.8.4       
 [4] compiler_3.5.1    pillar_1.3.0      git2r_0.23.0     
 [7] bindr_0.1.1       R.methodsS3_1.7.1 R.utils_2.7.0    
[10] tools_3.5.1       digest_0.6.17     lubridate_1.7.4  
[13] jsonlite_1.5      evaluate_0.11     nlme_3.1-137     
[16] gtable_0.2.0      lattice_0.20-35   pkgconfig_2.0.2  
[19] rlang_0.2.2       cli_1.0.1         rstudioapi_0.8   
[22] yaml_2.2.0        haven_1.1.2       withr_2.1.2      
[25] xml2_1.2.0        httr_1.3.1        knitr_1.20       
[28] hms_0.4.2         rprojroot_1.3-2   grid_3.5.1       
[31] tidyselect_0.2.4  glue_1.3.0        R6_2.3.0         
[34] readxl_1.1.0      rmarkdown_1.10    modelr_0.1.2     
[37] magrittr_1.5      whisker_0.3-2     backports_1.1.2  
[40] scales_1.0.0      htmltools_0.3.6   rvest_0.3.2      
[43] assertthat_0.2.0  colorspace_1.3-2  stringi_1.2.4    
[46] lazyeval_0.2.1    munsell_0.5.0     broom_0.5.0      
[49] crayon_1.3.4      R.oo_1.22.0