[BioC] intraSpotCorrelation consensus values for Single Channel analysis in limma

[Thierry Janssens]

Hello Jenny,

thank you for your swift reply.
Indeed, we use custom made 8x15k Agilent arrays made with 60mers from ESTs
from our non-model pet species. :)
Is this method then usable for the Agilent arrays ?

kind regards,

Thierry Janssens


> Hi Thierry,
>
> What kind of dual-channel arrays are these? I've seen very low
> intra-spot correlations from Agilent arrays, which have extremely
> high array to array spot consistency. Old-style pin-tip printed
> arrays had high intra-spot correlations because the amount of probe
> per spot could not be controlled very well from array to array.
>
> Cheers,
> Jenny
>
> At 10:16 AM 5/13/2009, Thierry Janssens wrote:
>>Dear BioC,
>>
>>while performing Single Channel Analysis in limma, according to
>>chapter 9 of the limma users guide, I notice that the R and G
>>foreground intensities are not correlated at all. I did not find a
>>thread about that problem on the forum. I am wondering what the
>>cause could be...
>>
>>The experiment is an unconnected/saturated design of 5 conditions,
>>on whcih I want to perform t-tests between the conditions.
>>
>>
>>...
>> > RGbc <- backgroundCorrect(RGlist, method = "edwards", offset = 30)
>> > MA <- normalizeWithinArrays(RGbc[j, ], method ="loess")
>> > targets
>>   Archive      Filename Cy5 Cy3
>>1     File  SSArray1.txt   A   B
>>2     File  SSArray2.txt   B   C
>>3     File  SSArray3.txt   C  AC
>>4     File  SSArray4.txt  AC  AB
>>5     File  SSArray5.txt  AB   A
>>6     File  SSArray6.txt   A   C
>>7     File  SSArray7.txt   C  AB
>>8     File  SSArray8.txt  AB   B
>>9     File  SSArray9.txt   B  AC
>>10    File SSArray10.txt  AC   A
>> > #sorteren op duplo
>> > o <- order(MA$genes$ProbeUID)
>> > MAsorted <- MA[o,]
>> > o <- order(MAbet$genes$ProbeUID)
>> > MAbetsorted <- MAbet[o,]
>> > r <- 0
>> > for(q in seq(1, nrow(MAbetsorted), 3)) {
>>+    r <- as.numeric((identical(MAbetsorted$genes$probeUID[q],
>>MAbetsorted$genes$probeUID[q+1]))
>>+    && (identical(MAbetsorted$genes$probeUID[q],
>>MAbetsorted$genes$probeUID[q+2])) ) + r
>>+ }
>> > r
>>[1] 5069
>> > # r moet 5069 zijn
>> > # Separate channel analysis in limma
>> > MAbetsortedav <- avedups(MAbetsorted, ndups = 3, spacing =1)
>> > targets <- readTargets("filelist.txt")
>> > targetstest <- targetsA2C(targets)
>> > u <- unique(targetstest$Target)
>> > f <- factor(targetstest$Target, levels=u)
>> > design <- model.matrix(~0+f)
>> > colnames(design) <- u
>> > design
>>   B A C AC AB
>>1  1 0 0  0  0
>>2  0 1 0  0  0
>>3  0 0 1  0  0
>>4  1 0 0  0  0
>>5  0 0 0  1  0
>>6  0 0 1  0  0
>>7  0 0 0  0  1
>>8  0 0 0  1  0
>>9  0 1 0  0  0
>>10 0 0 0  0  1
>>11 0 0 1  0  0
>>12 0 1 0  0  0
>>13 0 0 0  0  1
>>14 0 0 1  0  0
>>15 1 0 0  0  0
>>16 0 0 0  0  1
>>17 0 0 0  1  0
>>18 1 0 0  0  0
>>19 0 1 0  0  0
>>20 0 0 0  1  0
>>attr(,"assign")
>>[1] 1 1 1 1 1
>>attr(,"contrasts")
>>attr(,"contrasts")$f
>>[1] "contr.treatment"
>> > corfit <- intraspotCorrelation(MAbetsortedav, design)
>>Warning messages:
>>1: In remlscore(y, X, Z) : reml: Max iterations exceeded
>>2: In remlscore(y, X, Z) : reml: Max iterations exceeded
>> > corfit$consensus.correlation
>>[1] *0.06922669
>>
>>*In previous threads I read that this correlation should be 0.8-0.9
>>(after backtransformation with tanh). What now?
>>
>>
>>kind regards,
>>
>>Thierry
>>
>>--
>>Thierry K.S. Janssens
>>Vrije Universiteit Amsterdam
>>Faculty of Earth and Life Sciences
>>Institute of Ecological Science
>>Department of Animal Ecology,
>>De Boelelaan 1085
>>1081 HV AMSTERDAM, The Netherlands
>>Phone: +31 (0)20-5989147
>>Fax: +31 (0)20-5987123
>>thierry.janssens at ecology.falw.vu.nl
>>
>>
>>
>>_______________________________________________
>>Bioconductor mailing list
>>Bioconductor at stat.math.ethz.ch
>>https://stat.ethz.ch/mailman/listinfo/bioconductor
>>Search the archives:
>>http://news.gmane.org/gmane.science.biology.informatics.conductor
>
> Jenny Drnevich, Ph.D.
>
> Functional Genomics Bioinformatics Specialist
> W.M. Keck Center for Comparative and Functional Genomics
> Roy J. Carver Biotechnology Center
> University of Illinois, Urbana-Champaign
>
> 330 ERML
> 1201 W. Gregory Dr.
> Urbana, IL 61801
> USA
>
> ph: 217-244-7355
> fax: 217-265-5066
> e-mail: drnevich at illinois.edu
>
>