[BioC] Design Matrix: Dye-swap or channel-swap?

Gordon Smyth smyth at wehi.edu.au
Sat Dec 13 07:59:23 MET 2003

At 08:36 AM 13/12/2003, John Welsh wrote:
>Sorry for the formatting error.
>Mine is a two color experiment, where each of four chips is a biological
>replicate, and half of these are dye-swapped. The (biological) control is
>always in the first channel and the treatment is always in the other. My
>confusion must stem from the R/G mnenomic: Cy5 fluorescence is in the red
>and Cy3 in the green, but this is irrelevant. Limma's R/G refer to false

Limma's R/G corresponds to Cy5/Cy3 dyes where that information is 
available, for example with Genepix data. Otherwise "green" is a synonym 
for channel 1 and "red" is a synonym for channel 2. (If you break the 
convention that Cy3=channel 1 and Cy5=channel 2, then it's up to you to 
unravel it.)


>  My controls are always false colored red, and the experimentals
>always green, irrespective of the dye. Thus, all of the control values
>should go into one matrix, and all the experimental values into the other,
>and my design would be c(1,1,1,1). Half of the measurements in the R channel
>should be cy3 measurements and half should be cy5 measurements. That way,
>dye-biased measurements from the same spot, assuming no biological change,
>should average out to zero, after normalization. Does that sound right?
>John Welsh
>Associate Professor
>Sidney Kimmel Cancer Center
>10835 Altman Row
>San Diego, CA 92121
>(858) 450-5990 ex.282
>jwelsh at skcc.org

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