Hello all:
I am working on the RNA-Seq project on non-model organism. The genome is not available for this organism but the mitochondria has been sequenced and published.
I want to remove all mt sequence from my original fastq files so I am going to align my reads (100bp pair-end) to mt genome. I want to use Bowtie 2. But I found Bowtie 2 is designed for "quickly aligning large sets of short DNA sequences (reads) to large genomes".
The animal mt genome is relatively small so I am wondering if Bowtie 2 still work? I want to use this because one of its output option is unmapped reads, which is exactly what I want.
If it's RNA-Seq you should use a splice-aware aligner e.g. STAR, Tophat, ...
I totally missed the RNAseq part of this :(
One caveat about STAR is that you have to tweak setting to map against small genomes. I'm think this is mentioned in the documentation, but I saw a thread about this on SEQanswers this week so apparently this isn't quite documented well enough.
Generally, --genomeSAindexNbases needs to be scaled with the genome length, as ~min(14,log2(ReferenceLength)/2 - 1) at the genome index generation step.
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Nicolas Rosewick
A stupid question, but is there any splicing in the Mitochondria? Since it's more or less still a procaryote and the annotations I know don't include any junctions.
I happened to look this up and at least in some organisms the answer appears to be "yes, there's splicing".
Yeah, it'll still work fine. The test data set that comes with bowtie2 is from a phage lambda, whose genome is ~49kb (so not that much larger than a mitochondria).
Edit: D'OH, I missed that this is RNAseq! Follow NicoBxl 's advise!
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