How to align reads using bowtie against multiple customized reference files

Question

[Jackie]

Hi,

I am trying to use Bowtie to align RNA-seq fastq files to a set of small reference fasta files (i.e., ribosomal reference, mitochondrial reference, all for human). I've learned some groups have done that, but technically, I don't know how to implement that.

As far as I know, Bowtie does not directly align your fastq reads to the *.fasta files, you need to first build indexes using the *.fasta file(s), and then align the reads against the indexed reference. So, what I have now is a set of small reference fasta files, how can I directly align the reads against these multiple reference fasta files without indexing them first? Is that doable in bowtie?

Thank you!

Answer 1

[GenoMax]

Creating alignment indexes is an integral part of any NGS data alignment. One starts with the fasta files (multi-fasta in one single file is fine) and then creates the indexes. This is a one-time process. You can then re-use the indexes as needed.

You could use bbmap.sh from BBMap suite (Java program) if you don't want to pre-create the indexes and would rather do them on the fly. Details are in this thread.

You will need to pre-create the indexes with most other aligners.

If this is an effort to assess contamination from those kinds of reads then you could look at bbsplit.sh from BBMap suite to bin your reads using that same reference.