[BioC] HTqPCR setCategories problem

[Heidi Dvinge]

Hello Mike,

On 7 May 2010, at 17:11, Michael Muratet wrote:

> Greetings
>
> I have encountered a small problem when using the setCategories  
> method:
>
> s2010004660.cat <- setCategory(s2010004660, quantile=0.9,  
> groups=sampleNames(s2010004660))
> Categories after Ct.max and Ct.min filtering:
>              wt_4wks mt_4wks wt_17wks mt_17wks
> OK               582     571      566      578
> Undetermined     186     197      202      190
> Categories after standard deviation filtering:
>              wt_4wks mt_4wks wt_17wks mt_17wks
> OK               581     569      565      577
> Undetermined     186     197      202      190
> There were 50 or more warnings (use warnings() to see the first 50)
> > warnings()
> Warning messages:
> 1: In `[<-.factor`(`*tmp*`, index, value = "Unreliable") :
>   invalid factor level, NAs generated
> 2: In `[<-.factor`(`*tmp*`, index, value = "Unreliable") :
>   invalid factor level, NAs generated
> 3: In `[<-.factor`(`*tmp*`, index, value = "Unreliable") :
>   invalid factor level, NAs generated
>
Hm, I can't reproduce this with the standard data sets in HTqPCR,  
although this also only contains "OK" and "Undetermined" initially:

 > data(qPCRraw)
 > apply(featureCategory(qPCRraw), 2, table)
              sample1 sample2 sample3 sample4 sample5 sample6
OK               353     312     360     339     335     336
Undetermined      31      72      24      45      49      48
 > setCategory(qPCRraw, quantile=0.9, groups=rep(c("A", "B"), 3))
Categories after Ct.max and Ct.min filtering:
              sample1 sample2 sample3 sample4 sample5 sample6
OK               313     264     327     295     296     286
Undetermined      68     119      56      86      86      96
Unreliable         3       1       1       3       2       2
Categories after standard deviation filtering:
              sample1 sample2 sample3 sample4 sample5 sample6
OK               309     258     323     291     292     281
Undetermined      68     119      56      86      86      96
Unreliable         7       7       5       7       6       7

How does featureCategory() of your new object look after you run  
setCategory? Also, it seems like all your Ct values fall within the  
default range given by Ct.max and Ct.min in setCategory(), hence none  
of the categories are adjusted during the "first round", before the  
standard deviation filtering. What happens if you set one of Ct.max  
or Ct.min so that some values are called as "Unreliable" based on  
this, e.g. by saying Ct.max=25? Do you still get the same warning?

Cheers
\Heidi


> other 47 warnings are the same
>
> The categories that you get in the SDS data are "OK" and  
> "Undetermined" and it seems to be unwilling to add the new level  
> "Unreliable".  I tried to manually add the levels:
>
> featureCategory(s2010004660)$wt_4wks <- factor(featureCategory 
> (s2010004660)$wt_4wks, levels=c(levels(featureCategory(s2010004660) 
> $wt_4wks),"Unreliable"))
> featureCategory(s2010004660)$mt_4wks <- factor(featureCategory 
> (s2010004660)$mt_4wks, levels=c(levels(featureCategory(s2010004660) 
> $mt_4wks),"Unreliable"))
> featureCategory(s2010004660)$wt_17wks <- factor(featureCategory 
> (s2010004660)$wt_17wks, levels=c(levels(featureCategory(s2010004660) 
> $wt_17wks),"Unreliable"))
> featureCategory(s2010004660)$mt_17wks <- factor(featureCategory 
> (s2010004660)$mt_17wks, levels=c(levels(featureCategory(s2010004660) 
> $mt_17wks),"Unreliable"))
>
> and get another error
>
> Error in count[names(tab), i] <- tab : subscript out of bounds
>
> Is this a bug or operator error?
>
> Thanks
>
> Mike
>
> > sessionInfo()
> R version 2.11.0 (2010-04-22)
> i386-apple-darwin9.8.0
>
> locale:
> [1] en_US.UTF-8/en_US.UTF-8/C/C/en_US.UTF-8/en_US.UTF-8
>
> attached base packages:
> [1] stats     graphics  grDevices utils     datasets  methods   base
>
> other attached packages:
> [1] HTqPCR_1.2.0       limma_3.4.0        RColorBrewer_1.0-2  
> Biobase_2.8.0
>
> loaded via a namespace (and not attached):
> [1] affy_1.26.0           affyio_1.16.0          
> gdata_2.7.1           gplots_2.7.4          gtools_2.6.1           
> preprocessCore_1.10.0
> [7] tools_2.11.0
>
> Michael Muratet, Ph.D.
> Senior Scientist
> HudsonAlpha Institute for Biotechnology
> mmuratet at hudsonalpha.org
> (256) 327-0473 (p)
> (256) 327-0966 (f)
>
> Room 4005
> 601 Genome Way
> Huntsville, Alabama 35806
>
>
>
>