应该是收集了水稻所有的ChIP-seq数据的一个网站Plantseq(含ChIP-seq部分分析及可视化命令脚本)

Plantseq

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看看更新频率,你没看错,前几天才更新。。


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随便拿一个基因输入看看(注意这里只支持RAPDB-ID):Os11g0559200
  • 首先是这样的


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  • 选择右边的tracks(分析的人都知道,这一班就意味着各种data),分八个板块


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  • 比如我们想看组蛋白H3K27me3修饰,理所当然快捷选择antibody


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  • 然后我这里选择看了水稻野生型DJ里面12天叶片的H3K27me3修饰


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  • 可以看到这个基因上几乎没有H3K27me3修饰


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  • 特别Nice的是还可以支持导入自己的数据,像bw、gff文件等。


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其次特别Nice的是给了所有分析脚本

  • 链接:http://www.plantseq.org/tutorial
    • 使用的软件
      sratoolkit (v2.9.0)
      FastQC (v0.10.1)
      Bowtie2 (v2.3.4)
      samtools (v1.7)
      deeptools2

分析命令

$ ./fastq-dump -I SRRxxxxx (for single-end sequencing)
$ ./fastq-dump -I --split-files SRRxxxxx (for pair-end sequencing)
  • 使用软件版本以及质量过滤说明
    • Reads qualities were analyzed with FastQC (v0.10.1) and the low quality bases (less than 30) were ignored in the following mapping step.

    • All the raw reads were mapped to corresponding genomes with Bowtie2 (v2.3.4) and compressed to binary files using samtools (v1.7).

比对到基因组

$ bowtie2 -5 xx -3 xx -x /directory/tair10 -U file.fastq -S file.sam
  • Here, xx representsthe the position of the base will be trimmed from the reads before mapping. -5 and -3 indicates the direction of trimming (5’ end and 3’end, respectively); -x and tair10 indicate the mapping index. The index has to be built before mapping. Please see detail in bowtie2 manual.

将sam文件转换成bam文件

$ samtools view -bS file.sam > file.bam
$ samtools sort -T file -o file_sort.bam file.bam
$ samtools index file_sort.bam
  • Finally, the mapped reads were converted to bigwig format for visualization by using bamcoverage (deeptools2) with the following settings: binsize 10, normalizing with 1x genome content (RPGC, Arabidopsis thaliana, 135,000,000; Glycine max, 978,000,000; Oryza sativa, 375,000,000; Zea mays, 2,135,000,000), ignorDuplicates, samFlagExclude 1796 and 1924 for single and pair-end sequencing, respectively. The following command will convert single-end Arabidopsis thaliana sorted bam file to bigwig file.
$ bamCoverage -b flie_sort.bam -o file_sort.bw -bs 10 --effectiveGenomeSize 135000000 --normalizeUsing RPGC --ignoreDuplicates -e 100 --samFlagExclude 1796

2.1.3 Generating peak plateau, FRiP, and median enrichment

  • The following shell and three R scripts are used for generating figures of peak plateau, FRiP, and median enrichment from mapped binary file (bam file). A list of file name needs to be provided before running the script. All the files including scripts, bam files, and listshould be in the same directory.

scripts

  • 脚本内容


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Before running the script, several tools need to be installed and added to your executable path.

Bedtools
MACS2
R
ggplot2 package

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