<p>Hi everyone,</p>

<p>I'm having a bit of trouble finding out how to use fimo to discover TFBS. The questions I had were:</p>

<ol>
<li>How do you decide on good thresholds for PWM matching (--thresh)</li>
<li>How much do the backgrounds matter, should these be calculated from the sequences you're submitting? </li>
<li>Does how does Fimo handle masked fasta files? Is it better to submit hard of soft masked files?</li>
</ol>

<p>Any other tips on good usage for this program would be much appreciated as I'm finding the documentation quite vague.</p>

<p>My response is probably not that great but since nobody else is answering this question:</p>

<ol>
<li>you probably want to set a q-value of something like &lt;0.05 or &lt;0.01 since those cutoffs are pretty standard</li>
<li>backgrounds matter a lot, they should be calculated from the sequence that you are looking for motifs in.</li>
<li>how are you masking the files? with upper and lower cases or with X/Ns? you should be submitting sequence that you want to look for motifs in. If you are uninterested in motifs in repetitive regions, then you should mask them out (with Ns). </li>
</ol>

<p>Lastly, have you tried using <a href='http://biowhat.ucsd.edu/homer/'>HOMER</a>? The documentation on it is quite extensive and you might find it easier to use</p>
