RNAseq analysis

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In regards to RNAseq analysis:

I’m in the process of analysing RNA-seq data and I initially followed the HISAT2-StringTie-Ballgown pipeline, however, the most up to date version of ballgown does not recognize my StringTie files for some reason? It only recognizes tables from StringTiemerge, even tho the other StringTie outputs appear to be in the same format (tabular). I attempted an older version of Ballgown, which does recognize the StringTie files, however, it fails immediately after launching. I am now attempting to run Cuffquant to subsequently run Cuffdiff, thus trying a different pipeline, but I know the tools are deprecated and they are running very slowly.
I was hoping someone might have some tips regarding Ballgown, or could clarify if Cuffquant/Cuffdiff are still okay to use?

Asta

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I did not work with Ballgown but maybe this paper will be useful https://www.nature.com/articles/nprot.2016.095

As far as I know TopHat(2) is definitely outdated. TopHat uses a more simple approach based on empirical evidence in comparison to HISAT.

Cufflinks has not been maintained in years, however it looks like is still widely accepted (https://pdfs.semanticscholar.org/727b/ef807477c2e7fc2333328a19422fdebbe525.pdf)

You can look into salmon/kallisto/sailfish and the sleuth pipeline (https://pachterlab.github.io/sleuth_walkthroughs/trapnell/analysis.html) as an alternative to Tuxedo suite.

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This tool was wrapped by a community contributor and I’m not sure if it will work with Stringtie input (likely just Cufflinks).

As @Sammy notes, Tophat and all of the Cuff* tools are considered deprecated in Galaxy (the entire Tuxedo suite of tools). None of the wrappers have been updated for quite some time.

Current RNA-seq protocols can be found here:

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