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<h1 class="title toc-ignore">Creat heatplots with deeptools</h1>
<h4 class="author"><em>First Last</em></h4>
<h4 class="date"><em>YYYY-MM-DD</em></h4>

</div>


<!-- The file analysis/chunks.R contains chunks that define default settings
shared across the workflowr files. -->
<!-- Update knitr chunk options -->
<!-- Insert the date the file was last updated -->
<p><strong>Last updated:</strong> 2018-02-02</p>
<!-- Insert the code version (Git commit SHA1) if Git repository exists and R
 package git2r is installed -->
<p><strong>Code version:</strong> 01f846f</p>
<!-- Add your analysis here -->
<div id="deeptools" class="section level3">
<h3>Deeptools</h3>
<p>Deeptools is a way to make plots similar to genomation but they have enrichment plots and heatmaps together. Also this is in bash so I do not need to get the bam files into R.</p>
<div id="install" class="section level4">
<h4>Install</h4>
<p>I added the deeptools package to my net-seq conda environment.</p>
</div>
<div id="running" class="section level4">
<h4>Running:</h4>
<p>Step 1: Create bigwig coverage files with bamcoverage</p>
<ul>
<li>bamCoverage -b reads.bam -o coverage.bw</li>
</ul>
<p>Step 2: computeMatrix</p>
<ul>
<li><p>I will need my normalized bigwig reads and the bed interval file (in my case PAS clusters)</p></li>
<li><p>ex: computeMatrix scale-regions -S <biwig file(s)> -R <bed file> -b 1000 -a 1000 -out <compressed matrix name.gz></p></li>
<li><p>–skipZeros (option- not included in first try)</p></li>
</ul>
<p>Step 3: Plot heatmap</p>
<ul>
<li><p>required –matrixFile, -m (from the compute matrix), -out (file name to save image.png)</p></li>
<li><p>–sortRegions descending</p></li>
<li><p>–plotTitle, -T</p></li>
</ul>
<p>I will run this first on the hES 3’ seq becasue we expect heavy enrichment.</p>
<pre class="bash"><code>#!/bin/bash


#SBATCH --job-name=deeptools_hES
#SBATCH --time=8:00:00
#SBATCH --partition=gilad
#SBATCH --mem=16G
#SBATCH --mail-type=END


module load Anaconda3  

source activate net-seq  

bamCoverage -b /project2/gilad/briana/Lianoglou.data/hES.hg38.sorted.bam -o hES.cov.bw  

computeMatrix reference-point -S hES.cov.bw -R /project2/gilad/briana/apa_sites/clusters.hg38.bed -b 1000 -a 1000 -out hES.hg38.gz

plotHeatmap --sortRegions descend -m hES.hg38.gz  -out hES.hg38.apa.png 
</code></pre>
<p>This plot shows enrichement of the 3’ seq on called PAS sites. The axis are wrong.</p>
<div class="figure">
<img src="../data/hES.hg38.apa.png" alt="hES 3’ coverage at APA sites" />
<p class="caption">hES 3’ coverage at APA sites</p>
</div>
<p>I will make this plot excluding exons. I will use bedtools intersect to get APA sites not in exons.</p>
<p>I want A interects B -v. This will give me regions in A not in B. A is the cluster file. /project2/gilad/briana/apa_sites/clusters.hg38.bed and B is /project2/gilad/briana/apa_sites/deeptools/exons_hg38.bed</p>
<p>This bash file is /project2/gilad/briana/apa_sites/deeptools/heatmatrix_hES_apa_excludeExons.sh</p>
<pre class="bash"><code>#!/bin/bash


#SBATCH --job-name=deeptools_hES_noexon
#SBATCH --time=8:00:00
#SBATCH --partition=gilad
#SBATCH --mem=16G
#SBATCH --mail-type=END


module load Anaconda3  

source activate net-seq  

bedtools intersect -v -a /project2/gilad/briana/apa_sites/clusters.hg38.bed -b /project2/gilad/yangili/hg38_exons.bed &gt; ../clusters.hg38.noExons.bed 


#bamCoverage -b /project2/gilad/briana/Lianoglou.data/hES.hg38.sorted.bam -o hES.cov.bw
# dont need to redo this. The file exsists

computeMatrix reference-point -S hES.cov.bw -R /project2/gilad/briana/apa_sites/clusters.hg38.noExons.bed -b 1000 -a 1000 -out hES.hg38.noExons.gz

plotHeatmap --sortRegions descend -T &quot;hES 3` APA enrichment exclude exons&quot; -m hES.hg38.noExons.gz  -out hES.hg38.apa.noExons.png 
</code></pre>
<div class="figure">
<img src="../data/hES.hg38.apa.noExons.png" alt="hES 3’ coverage at APA sites not in exons" />
<p class="caption">hES 3’ coverage at APA sites not in exons</p>
</div>
</div>
</div>
<div id="session-information" class="section level2">
<h2>Session information</h2>
<!-- Insert the session information into the document -->
<pre class="r"><code>sessionInfo()</code></pre>
<pre><code>R version 3.4.2 (2017-09-28)
Platform: x86_64-apple-darwin15.6.0 (64-bit)
Running under: macOS Sierra 10.12.6

Matrix products: default
BLAS: /Library/Frameworks/R.framework/Versions/3.4/Resources/lib/libRblas.0.dylib
LAPACK: /Library/Frameworks/R.framework/Versions/3.4/Resources/lib/libRlapack.dylib

locale:
[1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     

loaded via a namespace (and not attached):
 [1] compiler_3.4.2  backports_1.1.2 magrittr_1.5    rprojroot_1.3-2
 [5] tools_3.4.2     htmltools_0.3.6 yaml_2.1.16     Rcpp_0.12.15   
 [9] stringi_1.1.6   rmarkdown_1.8.5 knitr_1.18      git2r_0.21.0   
[13] stringr_1.2.0   digest_0.6.14   evaluate_0.10.1</code></pre>
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