ChIP-Seq Top2 in Yeast

Introduction

This wiki page combines the two sequencing experiments of ChIP-Seq on yeast samples udring June / July 2017.

• ChIP-Seq Top2 peak-calling
• ChIP-Seq Top2 peak-calling E2

Sample quality

In the above previous analyses, we undertook some Duplicate removel as this showed up to be an issue in the FastQC quality files.

However, the effect of this removal was very low. In any case, the MACS2 program takes care of duplicates so samples were only trimmed for adapters and quality.

Alignment to the S288C Reference

Bam quality, no filtering

First Experiment

• ULS1_IP_E1
• ULS1_INP_E1
• WT_IP_E1
• WT_INP_E1

Second Experiment

• ULS1_IP_E2
• ULS1_INP_E2
• WT_T2_IP_E2
• WT_T2_INP_E2
• ULS1_T2_IP_E2
• ULS1_T2_INP_E2

Alignment to the new W303 Reference

Bam quality, no filtering

First Experiment

• ULS1_IP_E1
• ULS1_INP_E1
• WT_IP_E1
• WT_INP_E1

Second Experiment

• ULS1_IP_E2
• ULS1_INP_E2
• WT_T2_IP_E2
• WT_T2_INP_E2
• ULS1_T2_IP_E2
• ULS1_T2_INP_E2

Bam quality, filtering applied

The bam files were filtered both for mapping quality and low integrity reads in the following manner:

samtools view -b -F 1820 -q 48 <INBAMFILE> -o ${OUTBAMFILE> -@ <NUMTHREADS>

First Experiment

• ULS1_IP_E1
• ULS1_INP_E1
• WT_IP_E1
• WT_INP_E1

Second Experiment

• ULS1_IP_E2
• ULS1_INP_E2
• WT_T2_IP_E2
• WT_T2_INP_E2
• ULS1_T2_IP_E2
• ULS1_T2_INP_E2

Bam quality, 2nd filtering applied

This time bam files were filtered to MAPQ 28:

samtools view -b -F 1820 -q 28 <INBAMFILE> -o ${OUTBAMFILE> -@ <NUMTHREADS>

First Experiment

• ULS1_IP_E1
• ULS1_INP_E1
• WT_IP_E1
• WT_INP_E1

Second Experiment

• ULS1_IP_E2
• ULS1_INP_E2
• WT_T2_IP_E2
• WT_T2_INP_E2
• ULS1_T2_IP_E2
• ULS1_T2_INP_E2

Differential Peak-calling

Macs2 incorporated differential peak calling somewhat later into its set of tools. The idea is to take two conditions and compare their peaks.

The pipeline to follow is probably Macs2's recommended one at Differntial Binding Events