How do you find ballgown package for differential expression analysis?

Question

[seta]

Hi all friends,

I'm doing RNA-seq analysis for an organism with the sequenced genome, I want to do differential expression analysis at the transcript level and also examine alternative splicing events. Based on enter link description here, I used STAR (instead of HISAT) for mapping, then Stringtie, and now for ballgown, I'm not totally sure about it as the original paper has just 3 citations since 2014. Could you please tell me your experience with this package, how results are accurate and reliable in your hand? Please let me know if you have any suggestion for an alternative package?

Thanks

[Saeed]

Hi Seta,

I wonder, how found ballgown package for DE analysis and alternative splicing? I used hisat2 , stringtie and DESeq2 for DE analysis and I am going to use ballgown for alternative splicing. could you please give me your best suggestion (package) for alternative splicing?

Thanks

Answer 1

[andrew.j.skelton73]

Differential isoform expression is tricky. Have you considered Kallisto for quantification and it's counterpart Sleuth for the differential isoform expression? For gene level analyses I'd recommend the DESeq2 workflow.

[seta]

Thank you. The main goal is to examine alternative splicing events on DE transcript. Just for making sure, I used STAR for mapping reads to the reference genome and stringtie for assembly. Please kindly tell me if the resulted bam file can be accurately quantified by Kallisto? and Please let me know if the produced count by Kallisto can be fed to edgeR for differential expression transcript analysis?

[WouterDeCoster]

Kallisto operates on fastq files, not on bam files. For more information on kallisto, see the link in the post of andrew.j.skelton73. For import of counts from Kallisto/Salmon/... to edgeR/DESeq2/... you can use the tximport package.