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Copy pathChIPseq-NoSpikeIn.sh
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ChIPseq-NoSpikeIn.sh
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#!/bin/bash
#SBATCH --job-name=ChIP-seq_nospike
#SBATCH --ntasks=1
#SBATCH --mem=24G
#SBATCH -N 1
#SBATCH -n 16
#SBATCH --time=20:00:00
#SBATCH -e ChIP-seq_nospike.%job.err
#this script aligns reads to the mouse genome only, not a merged human/mouse genome. there is also no math to account for spike-in normalization. otherwise, it is the same as the spike-in script
module load bowtie/1.2.3
module load samtools
module load fastqc
Genome_DIR=/proj/seq/data/MM10_UCSC/Sequence/BowtieIndex/genome
chromSizes='/path/to/chromsizes/mm10.chrom.sizes'
RepeatSequence='/path/to/repeat/sequences/RepeatSequence.bed'
DIR=/path/to/fastqfiles/*fastq.gz
for f in ${DIR}
do
fastqc ${f}
(bowtie -v 2 -p 24 -S -m 1 --best --strata ${Genome_DIR} ${f} > ${f}.sam) 2>${f}.log
#The sam file must be converted to a sorted bam file.
samtools view -b -S -F 4 -@ 24 -o ${f}.bam ${f}.sam
samtools sort -@ 24 -o ${f}-sort.bam ${f}.bam
#Next, remove PCR duplicates.
samtools sort -n -@ 24 ${f}-sort.bam -o ${f}-sortedbyname.bam
samtools fixmate -m -@ 24 ${f}-sortedbyname.bam ${f}-fixmate.bam
samtools sort -@ 24 ${f}-fixmate.bam -o ${f}-fixmate_sorted.bam
samtools markdup -r -s -@ 24 ${f}-fixmate_sorted.bam ${f}-markdup.bam
samtools sort -@ 24 -o ${f}-markdup-sort.bam ${f}-markdup.bam
samtools index ${f}-markdup-sort.bam ${f}-markdup-sort.bam.bai
module load bedtools
#
##Here are the variables you need to change before running the script.FASTQ_FILE should be the unzipped fastq file.
#
bedtools bamtobed -i ${f}-markdup-sort.bam > ${f}.bed
awk '{
if ($6 == "-")
print $1, $2-200, $3, $4, $5, $6;
else
print $1, $2, $3+200, $4, $5, $6;
}' ${f}.bed > ${f}-sort-extend.bed
awk '{
if ($2 < 0)
print $1, 0, $3, $4, $5, $6;
else
print $1, $2, $3, $4, $5, $6;
}' ${f}-sort-extend.bed > ${f}-fixstart.bed
awk '{OFS="\t"}{
if (($1 == "chr1") &&
($3 > 195471971))
print $1, $2, 19541971, $4, $5, $6;
else if (($1 == "chr2") &&
($3 > 182113224))
print $1, $2, 182113224, $4, $5, $6;
else if (($1 == "chr3") &&
($3 > 160039680))
print $1, $2, 160039680, $4, $5, $6;
else if (($1 == "chr4") &&
($3 > 156508116))
print $1, $2, 156508116, $4, $5, $6;
else if (($1 == "chr5") &&
($3 > 151834684))
print $1, $2, 151834684, $4, $5, $6;
else if (($1 == "chr6") &&
($3 > 149736546))
print $1, $2, 149736546, $4, $5, $6;
else if (($1 == "chr7") &&
($3 > 145441459))
print $1, $2, 145441459, $4, $5, $6;
else if (($1 == "chr8") &&
($3 > 129401213))
print $1, $2, 129401213, $4, $5, $6;
else if (($1 == "chr9") &&
($3 > 124595110))
print $1, $2, 124595110, $4, $5, $6;
else if (($1 == "chr10") &&
($3 > 130694993))
print $1, $2, 130694993, $4, $5, $6;
else if (($1 == "chr11") &&
($3 > 122082543))
print $1, $2, 122082543, $4, $5, $6;
else if (($1 == "chr12") &&
($3 > 120129022))
print $1, $2, 120129022, $4, $5, $6;
else if (($1 == "chr13") &&
($3 > 120421639))
print $1, $2, 120421639, $4, $5, $6;
else if (($1 == "chr14") &&
($3 > 124902244))
print $1, $2, 124902244, $4, $5, $6;
else if (($1 == "chr15") &&
($3 > 104043685))
print $1, $2, 104043685, $4, $5, $6;
else if (($1 == "chr16") &&
($3 > 98207768))
print $1, $2, 98207768, $4, $5, $6;
else if (($1 == "chr17") &&
($3 > 94987271))
print $1, $2, 94987271, $4, $5, $6;
else if (($1 == "chr18") &&
($3 > 90702639))
print $1, $2, 90702639, $4, $5, $6;
else if (($1 == "chr19") &&
($3 > 61431566))
print $1, $2, 61431566, $4, $5, $6;
else if (($1 == "chrX") &&
($3 > 171031299))
print $1, $2, 171031299, $4, $5, $6;
else if (($1 == "chrY") &&
($3 > 91744698))
print $1, $2, 91744698, $4, $5, $6;
else if (($1 == "chrM") &&
($3 > 16299))
print $1, $2, 16299, $4, $5, $6;
else if (($1 == "chr5_JH584299_random") &&
($3 > 953012))
print $1, $2, 953012, $4, $5, $6;
else if (($1 == "chrX_GL456233_random") &&
($3 > 336933))
print $1, $2, 336933, $4, $5, $6;
else if (($1 == "chrY_JH584301_random") &&
($3 > 259875))
print $1, $2, 259875, $4, $5, $6;
else if (($1 == "chr1_GL456211_random") &&
($3 > 241735))
print $1, $2, 241735, $4, $5, $6;
else if (($1 == "chr4_GL456350_random") &&
($3 > 227966))
print $1, $2, 227966, $4, $5, $6;
else if (($1 == "chr4_JH584293_random") &&
($3 > 207968))
print $1, $2, 207968, $4, $5, $6;
else if (($1 == "chr1_GL456221_random") &&
($3 > 206961))
print $1, $2, 206961, $4, $5, $6;
else if (($1 == "chr5_JH584297_random") &&
($3 > 205776))
print $1, $2, 205776, $4, $5, $6;
else if (($1 == "chr5_JH584296_random") &&
($3 > 199368))
print $1, $2, 199368, $4, $5, $6;
else if (($1 == "chr5_GL456354_random") &&
($3 > 195993))
print $1, $2, 195993, $4, $5, $6;
else if (($1 == "chr4_JH584294_random") &&
($3 > 191905))
print $1, $2, 191905, $4, $5, $6;
else if (($1 == "chr5_JH584298_random") &&
($3 > 184189))
print $1, $2, 184189, $4, $5, $6;
else if (($1 == "chrY_JH584300_random") &&
($3 > 182347))
print $1, $2, 182347, $4, $5, $6;
else if (($1 == "chr7_GL456219_random") &&
($3 > 175968))
print $1, $2, 175968, $4, $5, $6;
else if (($1 == "chr1_GL456210_random") &&
($3 > 169725))
print $1, $2, 169725, $4, $5, $6;
else if (($1 == "chrY_JH584303_random") &&
($3 > 158099))
print $1, $2, 158099, $4, $5, $6;
else if (($1 == "chrY_JH584302_random") &&
($3 > 155838))
print $1, $2, 155838, $4, $5, $6;
else if (($1 == "chr1_GL456212_random") &&
($3 > 153618))
print $1, $2, 153618, $4, $5, $6;
else if (($1 == "chrUn_JH584304") &&
($3 > 114452))
print $1, $2, 114452, $4, $5, $6;
else if (($1 == "chrUn_GL456379") &&
($3 > 72385))
print $1, $2, 72385, $4, $5, $6;
else if (($1 == "chr4_GL456216_random") &&
($3 > 66673))
print $1, $2, 66673, $4, $5, $6;
else if (($1 == "chrUn_GL456393") &&
($3 > 55711))
print $1, $2, 55711, $4, $5, $6;
else if (($1 == "chrUn_GL456366") &&
($3 > 47073))
print $1, $2, 47073, $4, $5, $6;
else if (($1 == "chrUn_GL456367") &&
($3 > 42057))
print $1, $2, 42057, $4, $5, $6;
else if (($1 == "chrUn_GL456239") &&
($3 > 40056))
print $1, $2, 40056, $4, $5, $6;
else if (($1 == "chr1_GL456213_random") &&
($3 > 39340))
print $1, $2, 39340, $4, $5, $6;
else if (($1 == "chrUn_GL456383") &&
($3 > 38659))
print $1, $2, 38659, $4, $5, $6;
else if (($1 == "chrUn_GL456385") &&
($3 > 35240))
print $1, $2, 35240, $4, $5, $6;
else if (($1 == "chrUn_GL456360") &&
($3 > 31704))
print $1, $2, 31704, $4, $5, $6;
else if (($1 == "chrUn_GL456378") &&
($3 > 31602))
print $1, $2, 31602, $4, $5, $6;
else if (($1 == "chrUn_GL456389") &&
($3 > 28772))
print $1, $2, 28772, $4, $5, $6;
else if (($1 == "chrUn_GL456372") &&
($3 > 28664))
print $1, $2, 28664, $4, $5, $6;
else if (($1 == "chrUn_GL456370") &&
($3 > 26764))
print $1, $2, 26764, $4, $5, $6;
else if (($1 == "chrUn_GL456381") &&
($3 > 25871))
print $1, $2, 25871, $4, $5, $6;
else if (($1 == "chrUn_GL456387") &&
($3 > 24685))
print $1, $2, 24685, $4, $5, $6;
else if (($1 == "chrUn_GL456390") &&
($3 > 24668))
print $1, $2, 24668, $4, $5, $6;
else if (($1 == "chrUn_GL456394") &&
($3 > 24323))
print $1, $2, 24323, $4, $5, $6;
else if (($1 == "chrUn_GL456392") &&
($3 > 23629))
print $1, $2, 23629, $4, $5, $6;
else if (($1 == "chrUn_GL456382") &&
($3 > 23158))
print $1, $2, 23158, $4, $5, $6;
else if (($1 == "chrUn_GL456359") &&
($3 > 22974))
print $1, $2, 22974, $4, $5, $6;
else if (($1 == "chrUn_GL456396") &&
($3 > 21240))
print $1, $2, 21240, $4, $5, $6;
else if (($1 == "chrUn_GL456368") &&
($3 > 20208))
print $1, $2, 20208, $4, $5, $6;
else if (($1 == "chr4_JH584292_random") &&
($3 > 14945))
print $1, $2, 14945, $4, $5, $6;
else if (($1 == "chr4_JH584295_random") &&
($3 > 1976))
print $1, $2, 1976, $4, $5, $6;
else
print $1, $2, $3, $4, $5, $6;
}' ${f}-fixstart.bed > ${f}-fixall.bed
module load macs/2.2.7.1
module load bedtools/2.25.0
macs2 callpeak -g mm -q 0.01 -f BED -n ${f} -t ${f}-fixall.bed
awk 'BEGIN{OFS="\t"}{print $1, $2-50, $3+50, $4, $5
}' ${f}_summits.bed > ${f}-summits-exp50.bed
bedtools intersect -a ${f}-summits-exp50.bed -b ${RepeatSequence} -v -wa > ${f}-summits-exp50-noreps.bed
awk 'BEGIN{OFS="\t"}{print $1, $2+50, $3-50, $4, $5
}' ${f}-summits-exp50-noreps.bed > ${f}-summits-exp50-noreps-shrink.bed
bedtools genomecov -bga -g ${chromSizes} -i ${f}-fixall.bed > ${f}-fixall.bedgraph
sort -k1,1 -k2,2n ${f}-fixall.bedgraph > ${f}-sort.bedgraph
module load ucsctools
bedGraphToBigWig ${f}-sort.bedgraph ${chromSizes} ${f}.bw
done
## Remove intermediate files when done
#rm *fix* *xls *bedgraph *r *narrowPeak *bam *sam *align* *summits.bed *log *bai