[BioC] exclude flagged spots in limma

[J.delasHeras at ed.ac.uk]

Quoting Yolande Tra <yvtsma at rit.edu>:

> Hi Gordon,
>
> I have read some of your answers regarding this issue but it does not 
> answer mine. I have 5 separate image output files from Scanalyze (not 
> one of the output listed in Limma). For each file, one of the column 
> named FLAG contains the flagged values of 1 if bad and 0 if good. Can 
> you please help me, how can I exclude these spots from the analysis.
>
> The following commands helped me to read each file saved ast text 
> (tab delimiter)
>> filenames <- c("gp1.dat","gp2.dat","gp3.dat","gp4.dat","gp5.dat")
>> RG <- read.maimages(filenames,annotation="My_spot_labels",
> columns=list(Rf="CH1I",Gf="CH2I",Rb="CH1B",Gb="CH2B"))
>  These are the notation in Scanalyze.
>
> I have used one function in the archive and got the following error
>  > mywtfun <- function(exclude.flags=c(1,2,3)) function(obj) 1-(obj$Flag %in%
> + exclude.flags)
>> RG <- read.maimages(filenames,annotation="My_spot_labels", 
>> columns=list(Rf="CH1I",Gf="CH2I",Rb="CH1B",Gb="CH2B"), 
>> wt.fun=mywtfun(c(1)))
> Error in "[<-"(`*tmp*`, , i, value = numeric(0)) :
>         nothing to replace with
>
> Thank you so much for your help.
> Yolande

Hi Yolande,

you can add "manually" a component $weights to your RG object. This is 
merely a matrix with values between 0 and 1, with as many columns as 
there are slides, and as many rows as there are genes. So each column 
is a "flags" column for each slide.

You can use the read.table function to read the whole data file (output 
from Scanalyze) for each slide, and then simply pick the column 
containing the flags, and use them to build the matrix. Then just 
assign RG$weights<- yourmatrix, and you're rolling...

You can create your own flags from any other parameters you wish, as 
long as you end up with a number between 0 and 1, where 1 is full 
weight (good) and 0 is bad, and different degrees in between.

I hope this helps.

Jose

-- 
Dr. Jose I. de las Heras                      Email: J.delasHeras at ed.ac.uk
The Wellcome Trust Centre for Cell Biology    Phone: +44 (0)131 6513374
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