[BioC] DESeq2 transcript level

Michael Love michaelisaiahlove at gmail.com
Fri May 9 19:45:32 CEST 2014


hi Alicia,

In your previous email, you didn't mention that you had used software
for estimating isoform specific counts, so I pasted a link to Simon's
answer concerning isoform counts. If one can assign reads to isoforms
with high confidence then it would be fine to do testing with DESeq2.
However, this is typically a difficult task which involves uncertainty
in estimation and DESeq2 does not take into account any uncertainty in
these estimated counts. If you have 10 reads in the count matrix,
DESeq2 will take this to mean 10 reads were unambiguously assigned to
this feature. Through estimation steps, this could be 10 reads plus or
minus 10 reads. So the quality of the results will depend on the
quality of the input.

Mike

On Fri, May 9, 2014 at 11:11 AM, Alicia R. Pérez-Porro
<alicia.r.perezporro at gmail.com> wrote:
> Hi Mike,
>
> Thanks for your answer. I read the explanation from Simon and am still a
> little bit confuse, reading the edgeR manual I found this: "edgeR can be
> applied to di fferential expression at the gene, exon, transcript or tag
> level. In fact, read counts can be summarized by any genomic feature. edgeR
> analyses at the exon level are easily extended to detect di fferential
> splicing or isoform-specifi c dif ferential expression".
>
> So, my initial idea was to treat each isoform like a gene because for the
> approach that I am attempting right now I am not interested in differences
> in expression of isoforms belonging to the same gene. I did my de novo with
> Trinity, alignment using Bowtie and estimation of abundance with RSEM. I
> created a matrix with the RSEM counts (isoforms results) and I was planning
> on using it as input for my next step.
>
> I understand that even I want to treat each isoform as a gene to do it with
> DESeq or DESeq2 is not a good idea. I really don't know about edgeR and am
> also considering EBseq.
>
> Suggestions? Thoughts?
>
> Thank you in advance.
> Alicia
>
>
>
> --
> Alicia R. Pérez-Porro
> PhD candidate
>
> Giribet lab
> Department of Organismic and Evolutionary Biology
> MCZ labs
> Harvard University
> 26 Oxford St, Cambridge MA 02138
> phone: +1 617-496-5308
> fax: +1 617-495-5667
> www.oeb.harvard.edu/faculty/giribet/
>
> Department of Marine Ecology
> Center for Advanced Studies of Blanes (CEAB-CSIC)
> C/Accés Cala St. Francesc 14
> 17300 Blanes, Girona, SPAIN
> phone: +34 972 336 101
> fax: +34 972 337 806
> www.ceab.csic.es
>
>
> On Fri, May 9, 2014 at 8:07 AM, Michael Love <michaelisaiahlove at gmail.com>
> wrote:
>>
>> hi Alicia,
>>
>> DESeq2 is intended for gene-level analysis. here is some explanation from
>> Simon on the list as to why doing a count-based analysis at the
>> transcript/isoform level is problematic:
>>
>> https://stat.ethz.ch/pipermail/bioconductor/2012-February/043410.html
>>
>> if you are interested in looking for differential exon usage, you can
>> instead consider using the DEXSeq package:
>>
>> publication
>> http://www.ncbi.nlm.nih.gov/pubmed/22722343
>>
>> software:
>> http://bioconductor.org/packages/release/bioc/html/DEXSeq.html
>>
>> Mike
>>
>>
>>
>> On Thu, May 8, 2014 at 7:26 PM, Alicia R. Pérez-Porro
>> <alicia.r.perezporro at gmail.com> wrote:
>>>
>>> Dear Bioconductor users,
>>>
>>> I am working with differential expression at the transcript (isoform)
>>> level. I have 6 different conditions (2 replicates/condition).
>>>
>>> I want to know if I can use DESeq2 for that or if the package can only be
>>> used at the gene level.
>>>
>>> Thanks,
>>> Alicia
>>>
>>>
>>> --
>>> Alicia R. Pérez-Porro
>>> PhD candidate
>>>
>>> Giribet lab
>>> Department of Organismic and Evolutionary Biology
>>> MCZ labs
>>> Harvard University
>>> 26 Oxford St, Cambridge MA 02138
>>> phone: +1 617-496-5308
>>> fax: +1 617-495-5667
>>> www.oeb.harvard.edu/faculty/giribet/
>>>
>>> Department of Marine Ecology
>>> Center for Advanced Studies of Blanes (CEAB-CSIC)
>>> C/Accés Cala St. Francesc 14
>>> 17300 Blanes, Girona, SPAIN
>>> phone: +34 972 336 101
>>> fax: +34 972 337 806
>>> www.ceab.csic.es
>>>
>>>         [[alternative HTML version deleted]]
>>>
>>>
>>> _______________________________________________
>>> Bioconductor mailing list
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>>
>>
>



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