<div dir="ltr">Dear Yarden,<div><br></div><div>thanks a lot for the info.</div><div><br></div><div>I have since looked at some recent TopHat outputs. </div><div><br></div><div>Since we are following quite a complicated (read: "sophisticated" ;) mapping strategy (map against gen and trx, map the trx coordinates back to gen, merge files, keep best alignments and discard non-unique primary alignments), re-creating the SAM file was anyway required, and the conversion to the required output should be easy.<br>
</div><div><br></div><div class="gmail_extra">Thanks a lot for your help and enjoy your travels!</div><div class="gmail_extra"><br></div><div class="gmail_extra">Best,</div><div class="gmail_extra">Alex<br><br><br><br><div class="gmail_quote">
On Fri, Aug 16, 2013 at 8:29 PM, Yarden Katz <span dir="ltr"><<a href="mailto:yarden@mit.edu" target="_blank">yarden@mit.edu</a>></span> wrote:<br><blockquote class="gmail_quote" style="margin:0 0 0 .8ex;border-left:1px #ccc solid;padding-left:1ex">
Hi Alex,<br>
<br>
See comments below:<br>
<div class="im"><br>
On Aug 13, 2013, at 6:56 AM, Alexander Kanitz wrote:<br>
<br>
> Hi everyone,<br>
><br>
> I am planning to use MISO but I want to be flexible with the aligner I use (read: don't want to use Bowtie/TopHat).<br>
><br>
> Since I did not find any detailed specs regarding this issue, I have a few questions:<br>
> What does MISO need out of a SAM/BAM file? Specifically, which tags are considered and which are required (assuming all others will be ignored)?<br>
<br>
</div>MISO tries to not rely on too many binary SAM/BAM flags since as you imply many different aligners follow slightly different conventions.<br>
<br>
The main flags used as strandedness if you have stranded data and paired-ness/orientation of reads if you have paired-end data. There's a flag to MISO that can tell it to group read pairs by ID, e.g. assuming that the reads have the format:<br>
<br>
readX/1<br>
readX/2<br>
<br>
or something similar that makes it clear that they are both mates of the same sequenced molecule.<br>
<div class="im"><br>
> And most importantly: in what format (one line, separate lines, CIGAR string, MD tag, etc) do split/spliced alignments have to be represented?<br>
<br>
</div>CIGAR string. I can add a note to the manual clarifying this.<br>
<div class="im"><br>
<br>
><br>
> I would be very grateful for some help here and ideally a few lines of a MISO-compatible SAM file including some split/spliced alignments if possible (or a link to such a file).<br>
<br>
</div>Any BAM file produced with Tophat and Bowtie 1 should be an example of a MISO-accepted BAM, and we normally use Tophat around here though have users that use other aligners. I am traveling and don't have access handy now, but it should be easy to find.<br>
<br>
One thing to note: MISO ignores insertions and deletions as they do not match the annotation -- so any reads with insertion/deletions in their CIGAR strings will be discarded.<br>
<div class="im"><br>
><br>
> Thanks a lot!<br>
><br>
> Best,<br>
> Alex<br>
><br>
><br>
> --<br>
> Alexander Kanitz, Ph.D.<br>
><br>
> Winterthurerstrasse 358<br>
> 8057 Zurich<br>
> Switzerland<br>
><br>
> Phone: <a href="tel:%2B41%2076%20278%2039%2036" value="+41762783936">+41 76 278 39 36</a><br>
> Email: <a href="mailto:alexander.kanitz@alumni.ethz.ch">alexander.kanitz@alumni.ethz.ch</a><br>
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<br>
</blockquote></div><br><br clear="all"><div><br></div>-- <br>Alexander Kanitz, Ph.D.<br><br>Winterthurerstrasse 358<br>8057 Zurich<br>Switzerland<br><br>Phone: +41 76 278 39 36<br>Email: <a href="mailto:alexander.kanitz@alumni.ethz.ch" target="_blank">alexander.kanitz@alumni.ethz.ch</a><br>
</div></div>