1. ATAC-seq : Introduction and data background

Data background

In this workshop, we will analyse different genomic data generated from wild-type HCT116 colon cancer cell-lines. These data will comprise of genome-wide chromatin accessibility measurements using ATACseq, histone modification of H3K4me3 and H3K4me1 marks and CTCF transcription factor binding using ChIPseq.

H3K4me3 is known to mark active promoters around their TSS, H3K4me1 are known to mark enhancer regions and the transcription factor CTCF is known to mediate enhancer-promoter interactions. Also, genomic regions of activity (active promoter or enhancers) are expected to have an open chromatin landscape to allow the binding of regulatory factors. Thus, integration of ATACseq data and ChIPseq data (H3K4me3, H3K4me1 and CTCF) from the same cell-line will give us an understanding of its regulatory landscape.

Raw data will be downloaded and re-processed from ENCODE and the following data will be analyzed -

Sample	Experiment	Assay	Replicate	Source
HCT116	ATACseq	Chromatin accessibility - R1	1	ENCFF121EPT
HCT116	ATACseq	Chromatin accessibility - R2	1	ENCFF795EHY
HCT116	ATACseq	Chromatin accessibility - R1	2	ENCFF624DNH
HCT116	ATACseq	Chromatin accessibility - R2	2	ENCFF157PAR

Download data

NOTE: You don't need to download any data, all of the data has been pre-downloaded and saved in home/<username>/data/fastqdata. You will directly access these data during the practical training session.

ATACseq

We will download the fastq files (2 isogenic replicates) containing sequence reads (paired end) from an ATACseq experiment done on the HCT116 cell-line from the ENCODE database.

wget https://www.encodeproject.org/files/ENCFF121EPT/@@download/ENCFF121EPT.fastq.gz \
-O ATAC_Rep1_ENCFF121EPT.fastq_R1.gz ;
wget https://www.encodeproject.org/files/ENCFF795EHY/@@download/ENCFF795EHY.fastq.gz \
-O ATAC_Rep1_ENCFF795EHY.fastq_R2.gz ;
wget https://www.encodeproject.org/files/ENCFF624DNH/@@download/ENCFF624DNH.fastq.gz \
-O ATAC_Rep2_ENCFF624DNH.fastq_R1.gz ;
wget https://www.encodeproject.org/files/ENCFF157PAR/@@download/ENCFF157PAR.fastq.gz \
-O ATAC_Rep2_ENCFF157PAR.fastq_R2.gz

hg38 genome index

We will also download the precomputed hg38 genome index reference for allignment from iGenomes

wget http://igenomes.illumina.com.s3-website-us-east-1.amazonaws.com/Homo_sapiens/NCBI/GRCh38/Homo_sapiens_NCBI_GRCh38.tar.gz

Public datasets

Often one has to use publicly available datasets. These datasets are widely available through Gene Expression Omnibus - GEO and Array Express. Raw data fastq from human samples are usually deposited in The database of Genotypes and Phenotypes - dbGAP and European Genome-phenome Archive - EGA and are available under protected access. These data are usually in SRA format and an array of tools called sra-tools are available to manipulate these formats prior to regular analysis. We will not talk about these files formats in this workshop, but we want to make the participants aware that most of the publicly available raw ChIPseq and ATACseq data are in SRA format which needs to be converted to fastq format using SRA-tools.