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Peak calling

MACS2是被广泛使用的call peak工具，MACS3与MACS2算法差异不大，MACS2的命令基本可以直接迁移到MACS3。

  1. mkdir $wkdir/4_peakcalling
  2. cd $wkdir/4_peakcalling
  4. macs3 callpeak \
  5.   -g 119481543 \
  6.   -t clean_bams/ZAT6_ABA_rep1_final.bam \
  7.   -c clean_bams/Col0_mock_rep1_final.bam \
  8.   --name raw_peaks/ZAT6_ABA_rep1 \
  9.   --format BAM \
  10.   --keep-dup all \
  11.   --qvalue 0.075 \
  12.   --cutoff-analysis \
  13.   1>logs/ZAT6_ABA_rep1_callpeak_with_macs3.log 2>&1

Peak 过滤筛选

方法一

  1. read.table(file = "../ChIP-seq-narrow/raw_peaks/ZAT6_ABA_rep1_cutoff_analysis.txt", header = T)
  4. python scripts/plot_macs_cutoff_analysis.py \
  5.   raw_peaks/ZAT6_ABA_rep2_cutoff_analysis.txt \
  6.   raw_peaks/ZAT6_ABA_rep2_cutoff_analysis.pdf \
  7.   1>logs/ZAT6_ABA_rep2_plot_macs_cutoff_analysis.log 2>&1
  9. awk '$9>3' \
  10.   raw_peaks/ZAT6_ABA_rep1_peaks.narrowPeak \
  11.   > clean_peaks/cutoff/ZAT6_ABA_rep1_peaks.narrowPeak

方法二

  1. bedtools intersect \
  2.   -f 0.5 \ # 重叠百分比
  3.   -r \ # 相互重叠
  4.   -a clean_peaks/cutoff/ZAT6_ABA_rep1_peaks.narrowPeak \
  5.   -b clean_peaks/cutoff/ZAT6_ABA_rep2_peaks.narrowPeak \
  6.   -wa \ # 输出 A 文件
  7.   > clean_peaks/intersect/ZAT6_ABA_peaks.narrowPeak

方法三

  1. idr \
  2.   --samples \
  3.   raw_peaks/ZAT6_ABA_rep1_peaks.narrowPeak \
  4.   raw_peaks/ZAT6_ABA_rep2_peaks.narrowPeak \
  5.   --idr-threshold 0.05 \
  6.   --output-file clean_peaks/idr/ZAT6_ABA_true_rep_idr.txt \
  7.   --plot \
  8.   --input-file-type narrowPeak \
  9.   --rank p.value \
  10.   1>logs/ZAT6_ABA_select_peaks_by_idr.log 2>&1

Peak 差异分析

Peak 可视化

Peak 注释

ChIPseeker

参考

公众号 | YuLabSMU | 文章发表：Current Protocols | ChIPseeker R 包操作指南

公众号 | 小白鱼的生统笔记 | 使用R包ChIPseeker进行peaks关联的基因注释