How to create kallisto index that does not include ribosomal genes

Question

[bioinfo]

Hello,

I would like to create a kallisto index that contains coding and noncoding RNA but also exclude the ribosomal genes. I know I need the cdna and ncrna fasta files from the ensembl website initially. However, I am not sure how to remove the ribosomal genes from the reference. Is there an easy way to remove the ribosomal genes?

Thank you

[rpolicastro]

Did you want to remove both rRNA and ribosomal protein coding genes? Also, why did you want to remove these in the first place? It may cause some of these reads to be assigned to incorrect transcripts.

[bioinfo]

I would like to remove only the rRNA. We found out that there was an rRNA contamination from a probe we used so we would like to remove those transcripts from our data.

[swbarnes2]

Then you pick out the rRNA genes after aligning to everything. Removing things you know are present from the reference could make the alignment inaccurate.

Answer 1

[Trivas]

A more traditional way of doing what you want to do is to create an "rRNA database" that you map your reads against then take the unmapped reads and map them against your reference genome of interest.

[bioinfo]

How would I create the rRNA database? Would I use silva?

[Carambakaracho]

SortMeRNA could be an option, too (initially designed for metagenomic data)

[ATpoint]

Why would that be necessary? Using isolated databases always comes at the risk of false alignments. If you quantify against a reference that includes rRNA together with all the other annotated transcripts then you can later easily remove the rRNAs from your count matrix.