im analyzing a miRNA run from illumina genome analyzer. i received the data with the adapter already trimmed off, and im now at the point of aligning my reads against a reference.
when i try and align the reads using bwa's default params:
bwa aln ref.fasta read.fasta > read.sai
im getting <10% alignment (~90% of the reads are not getting aligned).
when i tried aligning using novoalign (that has the -s parameter to allow trimming) around 50% of the reads were aligned. this leads me to believe there is a parameter set in bwa that is more suitable for short RNA (including miRNA) alignment.
thoughts?
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written
by
Doctoroots
1 answer
Some details worth considering:
How long are your reads? How long are the miRNAs you are looking for?
I have also seen on my data that bwa reported fewer alignments than some other aligners (even blat and lastz) given the standard parameters.
There are some parameters worth a check:
-n NUM max #diff (int) or missing prob under 0.02 err rate (float) [0.04]
Try increasing -n
-o INT maximum number or fraction of gap opens [1]
Try increasing to 2 or 3
-l INT seed length [32]
the default 32 seems a little large to me especially if your query is short, try reducing towards 8 if that is possible
I would also try bowtie, bfast and maybe some others. For all the seed length parameter is important to increase sensitivity.
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written
by
Michael Dondrup
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