FIMO is giving me repeat hits

[Susan Luong]

Hello,

I'm using FIMO to scan 3'UTR sequences for RNA-binding protein motifs. I manually entered in several motif sequences (e.g. CPE 'TTTTAT'). FIMO does a good job of finding these hits, but I'm also getting several false positives that are simply a repeat of a hit already found. For instance, a correct hit will be 'TTTTAT' at position 414-419. However, I will get a 2nd hit for 'TTTTTAT' at position 413-419, when I never specified the 2nd sequence. Is there a way to increase the stringency of FIMO other than increasing the p-value threshold?

Thanks!
Susan

[CharlesEGrant]

When you type in a consensus sequences as a motif, FIMO converts it to a Position Weight Matrix (PWM). This means that matches may be overlapping, and need not be exact. This is usually desirable, since bindings sites may in fact overlap, and mismatches to the canonical binding site typically reduce the strength and likelihood of binding, but don't curtail it altogether.

 I will get a 2nd hit for 'TTTTTAT' at position 413-419, when I never specified the 2nd sequence

Your first match only contained 6 positions, and this one contains 7 positions so I suspect that it was an imperfect match to one of the other motifs you specified. FIMO lists which motif it was matching in the left-most column of the output.

Is there a way to increase the stringency of FIMO other than increasing the p-value threshold?

No, adjusting the p-value threshold is the only control on FIMO's stringency.

If you simply wish to scan for exact matches to a consensus sequence, the MEME Suite command line tools includes fasta-grep.