Differential isoform expression analyses
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4.7 years ago
rmf ★ 1.2k

For gene-level differential expression analysis, one can use gene ontology or such databases to obtain functions of genes and then cluster genes by function which helps to make sense of the differentially expressed genes and link it to fitness assays or functional work.

I have results from RSEM. I have a bunch of isoforms that go up and down. I was wondering what sort of interesting things I could do with that information. Are there databases for isoforms? full transcripts? My question is basically what are the potential downstream workflow after differential transcript/isoform expression analyses?

Thanks.

RNA-Seq isoform transcript expression • 2.1k views
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I don't know of any real databases that make use of isoform level information, however I do like seeing genes that are not differentially expressed between conditions, against their child isoforms that are differentially expressed. It acts as a nice illustration that we're missing things when analysing at the gene level.

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If you're interested in pre-mrna splicing, you can figure out what type of splicing events lead to the over- or under-expression of certain genes. Is there any evidence of aberrant splicing that leads of NMD? Or just simply summarize the isoforms at gene-level and do a functional cluster with GO or GSEA.

If you're in fact interested in splicing, instead of isoforms, I would personally focus more on the expressions of spliced junctions and/or exons.

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4.7 years ago
ssv.bio ▴ 190

A few... 1) Transcript switching 2) Cassette exons (if any) 3) Alternate splicing (if any) 4) Effect on structure or development (effect of point 1)

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4.0 years ago

If you are interested in isoform switches a potential tool could be IsoformSwitchAnalyzeR ( http://bioconductor.org/packages/IsoformSwitchAnalyzeR/ ) which directly parses RSEM (count) data output (also supports Cufflinks/Cuffdiff,Kallisto and Salmon) and enables identification of isoform switches with predicted functional consequences where the consequences can be chosen from a long list but includes protein domains etc.

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This tool seems promising and I would ask the OP to give it a try.

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4.7 years ago
Manvendra Singh ★ 2.1k

Differential Isoform expression is consequence of Alternate splicing If the overall expression of gene is not changing but isoform expression is differential then its the case isoform switch in your case.

you can perform gene set enrichment analysis to see if alternatively spliced transcripts are part of specific pathways

hth

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How do you do a gse with transcripts? Or did you mean to do the gse on the genes that the transcript is derived from?

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4.7 years ago
Satyajeet Khare ★ 1.6k

I would add alternative promoter usage and alternative polyadenylation to the list. However, whether changes in isoform expression suggest alternative RNA processing or not, is tricky as all isoforms of single gene need to analyzed for relative differential expression. Instead, you can just extract gene IDs from isoform IDs and perform differential expression analysis as usual.

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