[BioC] Ringo and dye swaps

[jbethune]

Dear all,

I'm using Ringo to analyse agilent tiling arrays. I have two arrays
measuring the same sample but the second one is dye-swapped.

The obivious solution is to create a target-file like this:

SlideNumber FileName     Species Tissue  Cy3     Cy5     Antibody
dosR1       array1.txt   m_bovis unspec  input   dosR    dosR
dosR2       array2.txt   m_bovis unspec  dosR    input   dosR

where the Cy3 and Cy5 values are exchanged for the second array.

But this exchange of Cy3 and Cy5 values does not have any effect on the
result (the values are the same and the plot of the dye-swapped array
seems to be upside-down in both cases).

I would very much appreciate if someone could tell me if this approach
with the target file is right so that I can search for the error
elsewhere.


If you are interested in further details, this is the relevant code:

dataDir <- "data/";

used_genome <- "m. bovis";
load( file = file.path( dataDir, "m_bovis_gff.RData" ) );

replicateAnalysis <- function( arrayfiles, targetfile ) {
	library( "Ringo" );
	RG <- read.maimages( arrayfiles, source="agilent" );
	RG$targets <- readTargets( targetfile );
	pA <- extractProbeAnno( RG, "agilent", genome=used_genome,
microarray=paste( "Agilent Array with species", used_genome ) );
	X <- preprocess( RG[ RG$genes$ControlType == 0, ], method="nimblegen",
idColumn="ProbeName" );
	sampleNames( X ) <- X$SlideNumber;
	smoothX <- computeRunningMedians( X, modColumn="Antibody",
winHalfSize=500, min.probes=3, probeAnno=pA );
	sampleNames( smoothX ) <- paste( sampleNames( X ), "smooth", sep="." );
	plot( seq( nrow( exprs( smoothX ) ) ), exprs( smoothX )[,1], col="red",
type="l", xlab="", ylab="" );
	if( length( arrayfiles ) == 2 ) {
		lines( seq( nrow( exprs( smoothX ) ) ), exprs( smoothX )[,2],
col="blue", type="l" );
	}
	return( smoothX );
}

hm <- c( file.path( dataDir, "array1.txt" ), file.path( dataDir,
"array2.txt" ) );
target_hm <- file.path( dataDir, "targets_11_1_replicates" );

hm_result <- replicateAnalysis( hm, target_hm );
title( "HM Design replicates", xlab="position", ylab="fold change" );
legend( "topright", c( "dosR1", "dosR2" ), fill=c("red", "blue"),
col=c("red", "blue" ) );