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davetang committed Mar 25, 2024
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Expand Up @@ -270,12 +270,12 @@ <h2 id="converting-a-sam-file-to-a-bam-file">Converting a SAM file to a BAM file
<p>Size of SAM file.</p>
<pre><code class="language-bash">ls -lh eg/ERR188273_chrX.sam
</code></pre>
<pre><code>## -rw-r--r-- 1 root root 321M Mar 25 07:18 eg/ERR188273_chrX.sam
<pre><code>## -rw-r--r-- 1 root root 321M Mar 25 07:28 eg/ERR188273_chrX.sam
</code></pre>
<p>Size of BAM file.</p>
<pre><code class="language-bash">ls -lh eg/ERR188273_chrX.bam
</code></pre>
<pre><code>## -rw-r--r-- 1 root root 67M Mar 25 07:16 eg/ERR188273_chrX.bam
<pre><code>## -rw-r--r-- 1 root root 67M Mar 25 07:27 eg/ERR188273_chrX.bam
</code></pre>
<p>We can use <code>head</code> to view a SAM file.</p>
<pre><code class="language-bash">head eg/ERR188273_chrX.sam
Expand Down Expand Up @@ -312,9 +312,9 @@ <h2 id="converting-a-bam-file-to-a-cram-file">Converting a BAM file to a CRAM fi

ls -lh eg/ERR188273_chrX.[sbcr]*am
</code></pre>
<pre><code>## -rw-r--r-- 1 root root 67M Mar 25 07:16 eg/ERR188273_chrX.bam
## -rw-r--r-- 1 root root 40M Mar 25 07:18 eg/ERR188273_chrX.cram
## -rw-r--r-- 1 root root 321M Mar 25 07:18 eg/ERR188273_chrX.sam
<pre><code>## -rw-r--r-- 1 root root 67M Mar 25 07:27 eg/ERR188273_chrX.bam
## -rw-r--r-- 1 root root 40M Mar 25 07:28 eg/ERR188273_chrX.cram
## -rw-r--r-- 1 root root 321M Mar 25 07:28 eg/ERR188273_chrX.sam
</code></pre>
<p>You can use <code>samtools view</code> to view a CRAM file just as you would for a
BAM file.</p>
Expand Down Expand Up @@ -343,27 +343,27 @@ <h2 id="sorting-a-sambam-file">Sorting a SAM/BAM file</h2>
ls -l eg/ERR188273_chrX.bam
ls -l eg/sorted.bam
</code></pre>
<pre><code>## -rw-r--r-- 1 root root 69983526 Mar 25 07:16 eg/ERR188273_chrX.bam
## -rw-r--r-- 1 root root 69983599 Mar 25 07:18 eg/sorted.bam
<pre><code>## -rw-r--r-- 1 root root 69983526 Mar 25 07:27 eg/ERR188273_chrX.bam
## -rw-r--r-- 1 root root 69983599 Mar 25 07:29 eg/sorted.bam
</code></pre>
<p>You should use use additional threads (if they are available) to speed
up sorting; to use four threads, use <code>-@ 4</code>.</p>
<p>Time taken using one thread (default).</p>
<pre><code class="language-bash">time samtools sort eg/ERR188273_chrX.sam -o eg/sorted.bam
</code></pre>
<pre><code>##
## real 0m8.755s
## user 0m8.460s
## sys 0m0.248s
## real 0m8.748s
## user 0m8.458s
## sys 0m0.244s
</code></pre>
<p>Time taken using four threads.</p>
<pre><code class="language-bash">time samtools sort -@ 4 eg/ERR188273_chrX.sam -o eg/sorted.bam
</code></pre>
<pre><code>## [bam_sort_core] merging from 0 files and 4 in-memory blocks...
##
## real 0m2.905s
## user 0m10.518s
## sys 0m0.505s
## real 0m2.896s
## user 0m10.513s
## sys 0m0.484s
</code></pre>
<p>Many of the SAMtools subtools can use additional threads, so make use of
them if you have the resources!</p>
Expand Down Expand Up @@ -736,26 +736,26 @@ <h2 id="comparing-bam-files">Comparing BAM files</h2>
<pre><code class="language-bash">samtools mpileup -s -f test_ref.fa aln_bwa.bam aln_mm.bam | head -20
</code></pre>
<pre><code>## [mpileup] 2 samples in 2 input files
## 1 4020 C 1 ^]. E ] 1 ^]. E ]
## 1 4021 T 1 . J ] 1 . J ]
## 1 4022 C 1 . J ] 1 . J ]
## 1 4023 T 1 . J ] 1 . J ]
## 1 4024 G 1 . J ] 1 . J ]
## 1 4025 T 1 . J ] 1 . J ]
## 1 4026 T 1 . J ] 1 . J ]
## 1 4027 A 1 . J ] 1 . J ]
## 1 4028 T 1 . J ] 1 . J ]
## 1 4029 A 1 . J ] 1 . J ]
## 1 4030 G 1 . J ] 1 . J ]
## 1 4031 C 1 . J ] 1 . J ]
## 1 4032 G 1 . J ] 1 . J ]
## 1 4033 G 1 . J ] 1 . J ]
## 1 4034 G 1 . J ] 1 . J ]
## 1 4035 A 1 . J ] 1 . J ]
## 1 4036 T 1 . J ] 1 . J ]
## 1 4037 T 1 . J ] 1 . J ]
## 1 4038 A 1 . J ] 1 . J ]
## 1 4039 C 1 . J ] 1 . J ]
## 1 8238 G 1 ^]. &gt; ] 1 ^]. &gt; ]
## 1 8239 G 1 . &gt; ] 1 . &gt; ]
## 1 8240 A 1 . J ] 1 . J ]
## 1 8241 C 1 . J ] 1 . J ]
## 1 8242 A 1 . J ] 1 . J ]
## 1 8243 C 1 . J ] 1 . J ]
## 1 8244 T 1 . J ] 1 . J ]
## 1 8245 G 1 . J ] 1 . J ]
## 1 8246 C 1 . J ] 1 . J ]
## 1 8247 G 1 . J ] 1 . J ]
## 1 8248 A 1 . J ] 1 . J ]
## 1 8249 C 1 . J ] 1 . J ]
## 1 8250 A 1 . J ] 1 . J ]
## 1 8251 G 1 . J ] 1 . J ]
## 1 8252 T 1 . J ] 1 . J ]
## 1 8253 G 1 . J ] 1 . J ]
## 1 8254 A 1 . J ] 1 . J ]
## 1 8255 G 1 . J ] 1 . J ]
## 1 8256 G 1 . J ] 1 . J ]
## 1 8257 G 1 . J ] 1 . J ]
</code></pre>
<p>Another approach is to use
<a href="https://deeptools.readthedocs.io/en/develop/">deepTools</a> and the
Expand Down Expand Up @@ -920,14 +920,30 @@ <h2 id="coverage">Coverage</h2>
calculate depth (and much faster than <code>samtools depth</code>) per base or
within a given window. The output is given in a BED file, where the
fourth column indicates the coverage.</p>
<pre><code class="language-bash">mosdepth ERR188275 eg/ERR188273_chrX.bam
<pre><code class="language-bash">mosdepth ERR188273 eg/ERR188273_chrX.bam
gunzip -c ERR188273.per-base.bed.gz | head
</code></pre>
<pre><code>## gzip: ERR188273.per-base.bed.gz: No such file or directory
<pre><code>## chrX 0 21648 0
## chrX 21648 21718 1
## chrX 21718 251270 0
## chrX 251270 251391 1
## chrX 251391 265950 0
## chrX 265950 266021 1
## chrX 266021 266096 2
## chrX 266096 269848 0
## chrX 269848 269923 1
## chrX 269923 270095 0
</code></pre>
<p><code>mosdepth</code> coverage.</p>
<pre><code class="language-bash">cat ERR188273.mosdepth.summary.txt
</code></pre>
<pre><code>## chrom length bases mean min max
## chrX 156040895 76303957 0.49 0 40804
## total 156040895 76303957 0.49 0 40804
</code></pre>
<p>Coverage in using a 500 bp window.</p>
<pre><code class="language-bash">mosdepth -n --fast-mode --by 500 ERR188275_500 eg/ERR188273_chrX.bam
gunzip -c ERR188275_500.regions.bed.gz | head
<pre><code class="language-bash">mosdepth -n --fast-mode --by 500 ERR188273_500 eg/ERR188273_chrX.bam
gunzip -c ERR188273_500.regions.bed.gz | head
</code></pre>
<pre><code>## chrX 0 500 0.00
## chrX 500 1000 0.00
Expand Down Expand Up @@ -985,5 +1001,5 @@ <h2 id="stargazers-over-time">Stargazers over time</h2>

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