[BioC] edgeR: Likelihood ratio test error

KJ Lim jinkeanlim at gmail.com
Tue Aug 7 15:58:10 CEST 2012


Dear edgeR users community,

Good day.

I'm analysing my RNA-Seq data which has 2 different genotypes (treeHS,
treeLS) and time of treatment (0H, 3H,
24H, 96H) with edgeR. I would like to learn the differential
expression of treeHS vs treeLS at specific time points i.e. 24H.

Thanks to Prof Gordon and his colleagues, the latest edgeR user's
guide (Chapter 3) is very useful for my case.

I encountered an error message when I carried out the likelihood ratio
test with either makeContrasts or coefficients approach. The error
message:

   > hl.contrasts <- makeContrasts(HvsL.00 = H.00H-L.00H, HvsL.03 =
(H.03H-H.00H)-(L.03H-L.00H),LvsH.00 = L.00H-H.00H,LvsH.03 =
(L.03H-L.00H)-(H.03H-H.00H), levels=hl.design)

  >  lrt.HvsL00 <- glmLRT(hl.fit, contrast=hl.contrasts[,"HvsL.00"])
  Error in array(x, c(length(x), 1L), if (!is.null(names(x))) list(names(x),  :
    dims [product 0] do not match the length of object [11]
  ---------------------------------------------------------------------------------------
  > colnames(hl.fit)
  [1] "H.00H" "H.03H" "H.24H" "H.96H" "L.00H" "L.03H" "L.24H" "L.96H"

 > lrt.HvsL00 <- glmLRT(hl.fit,contrast=c(1,0,0,0,-1,0,0,0))
  Error in array(x, c(length(x), 1L), if (!is.null(names(x))) list(names(x),  :
    dims [product 0] do not match the length of object [11]


Could the community kindly please light me on this matter? What
correction I should make in this case?

Thank you very much for your help.

Best regards,
KJ Lim

> sessionInfo()
R version 2.15.1 (2012-06-22)
Platform: i486-pc-linux-gnu (32-bit)

locale:
 [1] LC_CTYPE=en_US.utf8       LC_NUMERIC=C
 [3] LC_TIME=en_US.utf8        LC_COLLATE=en_US.utf8
 [5] LC_MONETARY=en_US.utf8    LC_MESSAGES=en_US.utf8
 [7] LC_PAPER=C                LC_NAME=C
 [9] LC_ADDRESS=C              LC_TELEPHONE=C
[11] LC_MEASUREMENT=en_US.utf8 LC_IDENTIFICATION=C

attached base packages:
[1] splines   stats     graphics  grDevices utils     datasets  methods
[8] base

other attached packages:
[1] edgeR_2.6.10 limma_3.12.1

loaded via a namespace (and not attached):
[1] tools_2.15.1



On 6 August 2012 10:11, Gordon K Smyth <smyth at wehi.edu.au> wrote:
> Dear KJ Lim,
>
> Well, your "tree" column has 16 different entries, a different entry for
> every library.  Naturally this causes a problem.
>
> So either you have no replication of any experimental condition, or you have
> mistakingly used different labels for the same condition.
>
> In your earlier email below, you claimed to have just two genotypes (tree=HS
> and tree=LS), but in your latest targets file you have 16 different
> genotypes.  I'm guessing that the genotypes are actually H and L, so the
> entries in the targets file should be just H and L.
>
> That's as much advice as I have time to give you, and I hope that others
> will help you further.  I feel that I've given you good general advice that
> is applicable to your experiment.
>
>
> Best wishes
> Gordon
>
> ---------------------------------------------
> Professor Gordon K Smyth,
> Bioinformatics Division,
> Walter and Eliza Hall Institute of Medical Research,
> 1G Royal Parade, Parkville, Vic 3052, Australia.
> Tel: (03) 9345 2326, Fax (03) 9347 0852,
> http://www.statsci.org/smyth
>
> On Mon, 6 Aug 2012, KJ Lim wrote:
>
>> Dear Prof Gordon,
>>
>> Good day. Thanks for you prompt replied.
>>
>> Below is the output of cbind(targets,Group=Group)
>>
>>> cbind(targets,Group=Group)
>>
>>                  files treat tree  X  Group
>> 1  ./rawData/HS0H01.txt   00H   H1 NA 00H.H1
>> 2  ./rawData/HS0H02.txt   00H   H2 NA 00H.H2
>> 3  ./rawData/HS3H01.txt   03H   H3 NA 03H.H3
>> 4  ./rawData/HS3H02.txt   03H   H4 NA 03H.H4
>> 5  ./rawData/HS1D01.txt   24H   H5 NA 24H.H5
>> 6  ./rawData/HS1D02.txt   24H   H6 NA 24H.H6
>> 7  ./rawData/HS4D01.txt   96H   H7 NA 96H.H7
>> 8  ./rawData/HS4D02.txt   96H   H8 NA 96H.H8
>> 9  ./rawData/LS0H01.txt   00H   L1 NA 00H.L1
>> 10 ./rawData/LS0H02.txt   00H   L2 NA 00H.L2
>> 11 ./rawData/LS3H01.txt   03H   L3 NA 03H.L3
>> 12 ./rawData/LS3H02.txt   03H   L4 NA 03H.L4
>> 13 ./rawData/LS1D01.txt   24H   L5 NA 24H.L5
>> 14 ./rawData/LS1D02.txt   24H   L6 NA 24H.L6
>> 15 ./rawData/LS4D01.txt   96H   L7 NA 96H.L7
>> 16 ./rawData/LS4D02.txt   96H   L8 NA 96H.L8
>>
>> Thank you for your time.
>>
>> Best regards,
>> KJ Lim
>>
>>
>> On 6 August 2012 07:33, Gordon K Smyth <smyth at wehi.edu.au> wrote:
>>>
>>> The error message is pretty self-explanatory.  Apparently your design
>>> matrix
>>> has as many columns as there are libraries, so there are no degrees of
>>> freedom left from which to estimate variability.  According to the code
>>> and
>>> output you have given, this message should be impossible, so you must
>>> have
>>> changed the data in some way that I cannot see.
>>>
>>> If you had shown the output from cbind(targets,Group=Group), then I might
>>> have been able to say something useful.
>>>
>>> Best wishes
>>> Gordon
>>>
>>>
>>> On Mon, 6 Aug 2012, KJ Lim wrote:
>>>
>>>> Dear Prof Gordon,
>>>>
>>>> Good day.
>>>>
>>>> Thanks for your time to update the edgeR User's Guide. It is useful.
>>>>
>>>> I combined all my experiment factors into one combined factor and
>>>> described the experiment matrix like the example in the User's guide:
>>>>
>>>> > Group <- factor(paste(targets$treat, targets$tree,sep="."))
>>>> > cbind(targets,Group=Group)
>>>> > hl.design <- model.matrix(~0+Group)
>>>>
>>>> But, I encountered an error when I performed the estimate dispersion
>>>> for my data
>>>>
>>>> > hl <- estimateGLMCommonDisp(hl, hl.design)
>>>>  Warning message:
>>>>  In estimateGLMCommonDisp.default(y = y$counts, design = design,  :
>>>>    No residual df: setting dispersion to NA
>>>>
>>>> I tried to figure out what I have done wrong, unfortunately, I have no
>>>> luck on that. Could you or the community kindly please light me for
>>>> this matter?
>>>>
>>>> Thank you very much for your time.
>>>>
>>>>> sessionInfo()
>>>>
>>>>
>>>> R version 2.15.1 (2012-06-22)
>>>> Platform: i486-pc-linux-gnu (32-bit)
>>>>
>>>> locale:
>>>> [1] LC_CTYPE=en_US.utf8       LC_NUMERIC=C
>>>> [3] LC_TIME=en_US.utf8        LC_COLLATE=en_US.utf8
>>>> [5] LC_MONETARY=en_US.utf8    LC_MESSAGES=en_US.utf8
>>>> [7] LC_PAPER=C                LC_NAME=C
>>>> [9] LC_ADDRESS=C              LC_TELEPHONE=C
>>>> [11] LC_MEASUREMENT=en_US.utf8 LC_IDENTIFICATION=C
>>>>
>>>> attached base packages:
>>>> [1] stats     graphics  grDevices utils     datasets  methods   base
>>>>
>>>> other attached packages:
>>>> [1] edgeR_2.6.10 limma_3.12.1
>>>>
>>>> loaded via a namespace (and not attached):
>>>> [1] tcltk_2.15.1 tools_2.15.1
>>>>
>>>> Best regards,
>>>> KJ Lim



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