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Applied Genomics

Downstream ChIP-seq Analysis

GitHub Classroom Assignment

Data

You’ve been given some data from a reanalysis of the paper Genome-wide maps of chromatin state in pluripotent and lineage-committed cells using the nf-core/chipseq pipeline.

You’ve also been given a file containing the MysteryFactorX regions.

Instructions

IGV

  1. Visualize the regions in IGV.
  2. Combine the MysteryFactorX Regions with Peaks for H3K4me3 and H3K27me3 from Mikkelsen ‘07. Reproduce Figure 1 (without the ChIP-chip data) to the best of your ability using IGV, including the MysteryFactorX.

Figure 1 Mikkelsen 07 Figure 1

If you're using IGV Desktop

Select mm10 as the genome, or download a fasta for just chr11 for mm10.

MEME-ChIP

  1. Perform motif analysis on the MysteryFactorX Regions. Add a image of the motif to your README.
  2. What TF might the MysteryFactorX be?

Example motif logo:

Example Logo

Running MEME-ChIP on chr1 only

If running MEME-ChIP is taking a long time, you might create a sequences fasta of only chr1.

Terminal window
wget https://hgdownload.cse.ucsc.edu/goldenPath/mm10/chromosomes/chr1.fa.gz
gunzip chr1.fa.gz
samtools faidx chr1.fa
wget https://huggingface.co/datasets/funlab/applied-genomics/resolve/main/chipseq/MysteryFactorX_ChIPseq_mm10.bed
sed -i '/^chr1\s/!d' MysteryFactorX_ChIPseq_mm10.bed
bedtools slop -g chr1.fa.fai -i MysteryFactorX_ChIPseq_mm10.bed -b 50 > extended.bed
bedtools getfasta -bed extended.bed -fi chr1.fa -fo sequences.fa

Bedtools

  1. How many MysteryFactorX regions overlap with H3K4me3 peaks?
  2. How many MysteryFactorX regions overlap with H3K27me3 peaks?

Feel free to use igv-notebook, the IGV web app, or the IGV desktop application! igv-reports will also work!

You may write a bash script, Jupyter Notebook, Snakemake, Galaxy or a combination of all of them. Just document how to reproduce your analysis!

Useful Docs

Hints

  • Search a Gene name that’s in the center of the figure to get the location of Figure 1
  • Read the Figure description to get the size of the window used in IGV