[BioC] DEXseq error on estimatelog2FoldChanges

[Fabrice Tourre]

Because the deep sequencing experiment is very expensive (at least to
some not so rich labs), we cannot do a biological replication. We try
to find some hints through RNA-seq, then do benchmark experiment to
valid the sequencing results.


On Tue, Feb 4, 2014 at 9:23 AM, Fabrice Tourre <fabrice.ciup at gmail.com> wrote:
> Alejandro,
>
> Thank you very much. Yes. I have only one control and one knockdown,
> without replication. Do you have any suggestions how can I get the
> differences in exon in two condition. Or different alternative
> splicing in two different condition. DESeq2 can do this?
>
> On Tue, Feb 4, 2014 at 9:18 AM, Alejandro Reyes <alejandro.reyes at embl.de> wrote:
>> Dear Fabrice Tourre,
>>
>> Thanks for your answers.  If I understood correctly, you have only one
>> control and one knockdown, without replication?  The errors are likely
>> because of this (sorry if the messages are not informative enough!).
>>
>> DEXSeq is designed to work with replicated data in order to distinguish
>> biological variation from differences between your different biological
>> conditions.  Unfortunately, if you have no replicates in your experiment you
>> won't be able to estimate variation across replicates,  therefore not be
>> able to test for differences in exon usage across biological conditions.
>> There are many papers (e.g. doi:10.1186/gb-2010-11-10-r106,
>> 10.1038/nprot.2013.099, 10.1101/gr.133744.111) and discussions (e.g. in
>> SeqAnswers) about why this is relevant.
>>
>> Best regards,
>> Alejandro
>>
>>
>>
>>
>>
>>
>>> I am analyzing mouse samples.
>>>
>>>> any( duplicated( paste( geneIDs(ecs), exonIDs(ecs), sep=":") ) )
>>>
>>> [1] FALSE
>>>
>>>> any( duplicated( featureNames(ecs)) )
>>>
>>> [1] FALSE
>>>
>>>> design(ecs)
>>>
>>>
>>>                               countFile
>>>
>>> ~/projects/seqdata/data/tophat2/sample1/sample1.counts
>>> ~/projects/seqdata/data/tophat2/sample1/sample1.counts
>>>
>>> ~/projects/seqdata/data/tophat2/sample2/sample2.counts
>>> ~/projects/seqdata/data/tophat2/sample2/sample2.counts
>>>
>>>                                                         condition
>>> libType
>>>
>>> ~/projects/seqdata/data/tophat2/sample1/sample1.counts   control
>>> paired-end
>>>
>>> ~/projects/seqdata/data/tophat2/sample2/sample2.counts knockdown
>>> paired-end
>>>
>>> On Tue, Feb 4, 2014 at 3:39 AM, Alejandro Reyes <alejandro.reyes at embl.de>
>>> wrote:
>>>>
>>>> Dear Fabrice Tourre,
>>>>
>>>> Thanks for your interest in DEXSeq and for the report!
>>>> Some questions to get to the details of the warnings and
>>>> the error message:
>>>>
>>>> 1. what is the output of doing:
>>>>
>>>> any( duplicated( paste( geneIDs(ecs), exonIDs(ecs), sep=":") ) )
>>>> any( duplicated( featureNames(ecs)) )
>>>>
>>>> 2. What is the output of design(ecs) of your ExonCountSet object?
>>>>
>>>> Best regards,
>>>> Alejandro
>>>>
>>>>
>>>>
>>>>> Dear expert,
>>>>>
>>>>> When I use DEXSeq to find different expressed exon, I got this error.
>>>>> In my analysis, I only have two samples, one is case, one is control.
>>>>>
>>>>> ecs <- estimatelog2FoldChanges( ecs )
>>>>> Error in `row.names<-.data.frame`(`*tmp*`, value = c("geneID", "exonID",
>>>>> :
>>>>>     duplicate 'row.names' are not allowed
>>>>> In addition: There were 50 or more warnings (use warnings() to see the
>>>>> first 50)
>>>>>
>>>>>
>>>>> warnings()
>>>>> Warning messages:
>>>>> 1: In chol.default(XVX + lambda * I, pivot = TRUE) :
>>>>>     the matrix is either rank-deficient or indefinite
>>>>>
>>>>>
>>>>>
>>>>> sessionInfo()
>>>>> R version 3.0.0 (2013-04-03)
>>>>> Platform: x86_64-unknown-linux-gnu (64-bit)
>>>>>
>>>>> locale:
>>>>>    [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C
>>>>>    [3] LC_TIME=en_US.UTF-8        LC_COLLATE=en_US.UTF-8
>>>>>    [5] LC_MONETARY=en_US.UTF-8    LC_MESSAGES=en_US.UTF-8
>>>>>    [7] LC_PAPER=C                 LC_NAME=C
>>>>>    [9] LC_ADDRESS=C               LC_TELEPHONE=C
>>>>> [11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C
>>>>>
>>>>> attached base packages:
>>>>> [1] parallel  stats     graphics  grDevices utils     datasets  methods
>>>>> [8] base
>>>>>
>>>>> other attached packages:
>>>>> [1] DEXSeq_1.8.0       Biobase_2.22.0     BiocGenerics_0.8.0
>>>>>
>>>>> loaded via a namespace (and not attached):
>>>>>    [1] biomaRt_2.18.0       Biostrings_2.30.1    bitops_1.0-6
>>>>>    [4] GenomicRanges_1.14.4 hwriter_1.3          IRanges_1.20.6
>>>>>    [7] RCurl_1.95-4.1       Rsamtools_1.14.2     statmod_1.4.18
>>>>> [10] stats4_3.0.0         stringr_0.6.2        XML_3.98-1.1
>>>>> [13] XVector_0.2.0        zlibbioc_1.8.0
>>>>>
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>>>>>
>>