/usr/share/doc/last-align/last-pair-probs.html

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<body>
<div class="document" id="last-pair-probs">
<h1 class="title">last-pair-probs</h1>

<p>This program reads candidate alignments of paired DNA reads to a
genome, and:</p>
<ol class="arabic">
<li><p class="first">estimates the distribution of distances between paired reads,</p>
</li>
<li><p class="first">estimates the probability that each alignment represents the
genomic source of the read.</p>
</li>
</ol>
<p>&quot;Paired&quot; means that the reads come from either end of a DNA fragment,
in tail-to-tail orientation:</p>
<pre class="literal-block">
5' ----------&gt;................................. 3'
3' .................................&lt;---------- 5'
</pre>
<p>Or head-to-head orientation:</p>
<pre class="literal-block">
5' .................................----------&gt; 3'
3' &lt;----------................................. 5'
</pre>
<p>The program writes the alignments with &quot;mismap&quot; probabilities,
i.e. the probability that the alignment does not represent the genomic
source of the read. By default, it discards alignments with mismap
probability &gt; 0.01.</p>
<div class="section" id="simple-usage">
<h2>Simple usage</h2>
<p>Suppose we have paired DNA reads in a file called &quot;interleaved.fastq&quot;
(in fastq-sanger format), where the first two reads are paired, the
next two reads are paired, and so on.  We can align them to the human
genome like this:</p>
<pre class="literal-block">
lastdb -uNEAR hg human-genome.fasta
lastal -Q1 -e120 -i1 hg interleaved.fastq &gt; temp.maf
last-pair-probs temp.maf &gt; out.maf
</pre>
<p>Suppose we have paired reads in two files, where the two first reads
are paired, the two second reads are paired, and so on.  We can
interleave them like this:</p>
<pre class="literal-block">
fastq-interleave x.fastq y.fastq | lastal -Q1 -e120 -i1 hg &gt; temp.maf
</pre>
</div>
<div class="section" id="reads-from-potentially-spliced-rna-molecules">
<h2>Reads from potentially-spliced RNA molecules</h2>
<p>Use the -r option:</p>
<pre class="literal-block">
last-pair-probs -r temp.maf &gt; out.maf
</pre>
<p>Without -r, it assumes the distances between paired reads follow a
normal distribution.  With -r, it assumes the distances follow a
skewed (log-normal) distribution, which is much more appropriate for
spliced RNA.</p>
</div>
<div class="section" id="efficient-usage">
<h2>Efficient usage</h2>
<p>The preceding recipes make a potentially-huge temp file, and
last-pair-probs reads it twice: first to estimate the distance
distribution, and then to estimate alignment probabilities.  It is
more efficient to estimate the distance distribution from a small
sample of the data:</p>
<pre class="literal-block">
lastal -Q1 -e120 -i1 hg sample.fastq | last-pair-probs -e
</pre>
<p>Suppose this tells us that the mean distance is 250 and the standard
deviation is 38.5.  We can use that to estimate the alignment
probabilities:</p>
<pre class="literal-block">
lastal -Q1 -e120 -i1 hg all.fastq | last-pair-probs -f250 -s38.5 &gt; out.maf
</pre>
</div>
<div class="section" id="going-faster-by-parallelization">
<h2>Going faster by parallelization</h2>
<p>This will run the pipeline on all your CPU cores:</p>
<pre class="literal-block">
fastq-interleave x.fastq y.fastq |
parallel-fastq &quot;lastal -Q1 -e120 -i1 hg | last-pair-probs -f250 -s38.5&quot; &gt; out.maf
</pre>
<p>It requires GNU parallel to be installed
(<a class="reference external" href="http://www.gnu.org/software/parallel/">http://www.gnu.org/software/parallel/</a>).</p>
</div>
<div class="section" id="details">
<h2>Details</h2>
<ul>
<li><p class="first">The &quot;distance&quot; between a pair of reads means the distance between
their 5' ends.  Positive distance indicates tail-to-tail
orientation, and negative distance indicates head-to-head
orientation.  Negative distances are not considered when -r is used,
nor for circular chromosomes.</p>
</li>
<li><p class="first">The program reads one batch of alignments at a time (by looking for
lines starting with &quot;# batch&quot;).  It assumes there is exactly one DNA
read per batch: if it finds more than one, it will complain.  The
lastal -i1 option ensures there is one query per batch.</p>
</li>
<li><p class="first">The alignments may be in either format produced by lastal (maf or
tabular).  They must include header lines (of the kind produced by
lastal) describing the alignment parameters.</p>
</li>
<li><p class="first">If a read name ends in neither &quot;/1&quot; nor &quot;/2&quot;, the program appends
&quot;/1&quot; if it is the 1st in a pair or &quot;/2&quot; if it is the 2nd.</p>
</li>
<li><p class="first">It is also possible to supply the alignments in two files:</p>
<pre class="literal-block">
lastal -Q1 -e120 -i1 hg x.fastq &gt; temp1.maf
lastal -Q1 -e120 -i1 hg y.fastq &gt; temp2.maf
last-pair-probs temp1.maf temp2.maf &gt; out.maf
</pre>
</li>
</ul>
</div>
<div class="section" id="options">
<h2>Options</h2>
<blockquote>
<table class="docutils option-list" frame="void" rules="none">
<col class="option" />
<col class="description" />
<tbody valign="top">
<tr><td class="option-group">
<kbd><span class="option">-h</span>, <span class="option">--help</span></kbd></td>
<td>Print a help message and exit.</td></tr>
<tr><td class="option-group">
<kbd><span class="option">-r</span>, <span class="option">--rna</span></kbd></td>
<td>Specifies that the fragments are from potentially-spliced RNA.</td></tr>
<tr><td class="option-group">
<kbd><span class="option">-e</span>, <span class="option">--estdist</span></kbd></td>
<td>Just estimate the distribution of distances.</td></tr>
<tr><td class="option-group">
<kbd><span class="option">-m <var>M</var></span>, <span class="option">--mismap=<var>M</var></span></kbd></td>
<td>Don't write alignments with mismap probability &gt; M.</td></tr>
<tr><td class="option-group">
<kbd><span class="option">-f <var>BP</var></span>, <span class="option">--fraglen=<var>BP</var></span></kbd></td>
<td>The mean distance in bp.  (With -r, the mean of
ln[distance].)  If this is not specified, the program will
estimate it from the alignments.</td></tr>
<tr><td class="option-group">
<kbd><span class="option">-s <var>BP</var></span>, <span class="option">--sdev=<var>BP</var></span></kbd></td>
<td>The standard deviation of distance in bp.  (With -r, the
standard deviation of ln[distance].)  If this is not
specified, the program will estimate it from the alignments.</td></tr>
<tr><td class="option-group">
<kbd><span class="option">-d <var>PROB</var></span>, <span class="option">--disjoint=<var>PROB</var></span></kbd></td>
<td>The prior probability that a pair of reads comes from
disjoint locations (e.g., different chromosomes).  This may
arise from real differences between the genome and the source
of the reads, or from errors in obtaining the reads or the
genome sequence.</td></tr>
<tr><td class="option-group">
<kbd><span class="option">-c <var>CHROM</var></span>, <span class="option">--circular=<var>CHROM</var></span></kbd></td>
<td>Specifies that the chromosome named CHROM is circular.  You
can use this option more than once (e.g., -c chrM -c chrP).
As a special case, &quot;.&quot; means all chromosomes are circular.
If this option is not used, &quot;chrM&quot; is assumed to be circular
(but if it is used, only the specified CHROMs are assumed to
be circular.)</td></tr>
<tr><td class="option-group">
<kbd><span class="option">-V</span>, <span class="option">--version</span></kbd></td>
<td>Show version information and exit.</td></tr>
</tbody>
</table>
</blockquote>
</div>
<div class="section" id="tips">
<h2>Tips</h2>
<ul>
<li><p class="first">To go faster, try gapless alignment (add -j1 to the lastal options).
Often, this is only minusculely less accurate than gapped alignment.</p>
</li>
</ul>
</div>
<div class="section" id="reference">
<h2>Reference</h2>
<p>For more information, please see this article:</p>
<blockquote>
An approximate Bayesian approach for mapping paired-end DNA reads to
a reference genome.  Shrestha AM, Frith MC.  Bioinformatics 2013
29(8):965-972.</blockquote>
</div>
</div>
</body>
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last-align 712-1ubuntu1 / usr / share / doc / last-align / last-pair-probs.html
