Asking for unmapped genes in RNA sequencing via Galaxy

Question

[Atefeh]

Hi. I have a RNA sequencing dataset which analyzed with Galaxy usage and type of data is paired-end. The dataset has a good quality leading to ignore the trimming level. However, after mapping with STAR tool the uniquely mapped genes were just around 6%.The mapped to loci was 3.2% and unmapped due to short reads is 90% what should I do?

[GenoMax]

Please ask galaxy related questions on their help forum: https://help.galaxyproject.org/ They will have access to your data and can help you directly.

That said, are you using the right genome? Remember with STAR you will need to align to genome reference, not transcriptome. You can take a sample of your reads and blast them at NCBI to make sure all reads identify with the genome you expect them to be from.

[Atefeh]

Thank you. I will recheck it.

Answer 1

[swbarnes2]

Note that for STAR "unmapped due to short read" really just means "didn't map". So either your reads are mostly trash, or you are aligning to the wrong reference. And yeah, aligning to transcriptome is a mistake, STAR wants genome + matching gtf. But I would check that you have the right species.

[Atefeh]

thank you.