featureCounts to analyse miR RNA-seq data

mirna
featurecounts
gft
gff

#1

Hi there:

I am writing to ask your help in regard to use featureCounts to analyse miR RNA-seq data in https://usegalaxy.eu/. I have tried to use a gene annotation file mmu.gff3 that I previously downloaded from miRBase. However, I always get the same error:

Fatal error: Exit code 255 ()

I guess I must select some option from Advance options panel, but I can not find which one is. Would you be so kind to help me with this issue?

Thanks in advance


#2

Hello, Featurecounts expects GTF as the input format and will fail with GFF3.

The tool Convert Formats >> gffread might be an option. The tool is deprecated at https://usegalaxy.org but still active at https://usegalaxy.eu. You’ll need to carefully inspect the output and possibly make other changes to get the data into a standardized GTF format that meets the other criteria (many advanced options are specifying what features to count on, gene/transcript attribute content/format – and your data needs to match up).

GTF formatting and related data manipulation help:

You also might want to consider using a public Galaxy server that is domain-specific for miRNA analysis. Please see this archived Galaxy Biostars post for details and links: https://biostar.usegalaxy.org/p/28003/index.html

Contacting the MiRBase authors is also an option – they could have alternative advice for obtaining the annotation data in GTF format. Please find the contact information at the bottom of this page: http://www.mirbase.org/ftp.shtml

All (known!) Public Galaxy servers: https://galaxyproject.org/use/

The EU team may post back with additional suggestions. Thanks!