ChIP Seq Project

Genomics Course, Fall 2020

Examine genomic localization of twist, snail, and CTBP in developing drosophila embryos.

Objective: Perform ChIP Seq analysis to identify site of binding for three developmental transcription factors in drosophila.

Sample Table

alias	target	genotype	batch	time	type	Qscale	ResultFile
twi_ip_1	twist	-	1	2to4h	ip	solexa1.3	twist_ip.fastq.gz
sna_ip_1	snail	-	1	2to4h	ip	solexa1.3	snail_ip.fastq.gz
twisna_wce_1	-	-	1	2to4h	wce	solexa1.3	snail_twist_wce.fastq.gz
dl_ip_1	Dl	-	2	2to4h	ip	Illumina-1.8	Dme_Dl_Tl10b_2to4h_1_ip.fastq.gz
dl_ip_2	Dl	-	2	2to4h	ip	Illumina-1.8	Dme_Dl_Tl10b_2to4h_2_ip.fastq.gz
dl_wce_1	-	-	2	2to4h	wce	Illumina-1.8	Dme_Tl10b_2to4h_1_wce.fastq.gz

Overall Plan:

Align FASTQ files to drosophila genome using bowtie to produce an alignment in BAM format.
Use MACS to identify peaks in each data set.

Optional: Visualize reads in a genome browser Create coverage track and visualize in genome browser

# example of running bowtie to align fastq files to genome
# create a variable to point to our alignment index
INDEX=/n/data1/genomes/bowtie-index/dm6/dm6
gunzip -c twist_ip.fastq.gz | bowtie --best --strata -p 3 -n 2 -m 1 $INDEX --sam - | samtools view -S -b -o twist_ip.bam -
gunzip -c snail_twist_wce.fastq.gz | bowtie --best --strata -p 3 -n 2 -m 1 $INDEX --sam - | samtools view -S -b –o \
      snail_twist_wce.bam -

# bam files have to be sorted for macs
# sort the bam file
samtools sort twist_ip.bam -T tmp -o twist_ip.bam

# run macs to find peaks
macs2 callpeak -t twist_ip.bam -c snail_twist_wce.bam --name=twist --format=BAM -g dm -q 0.01 –no-model

Tips: there is a script in the fastq directory that will sort and index all the bam files in a directory. Put the script into the same directory as the BAM files, and run it there.

If you’re having problems with the links to the fastq files in the fastq directory, you can create your own local links easily. Copy the link_files.sh script, and run it in your local directory and it will create links there.