Hi all, my experiment is a 2-color direct comparison wt vs mutant, but repeated on 5 successive timepoints. each timepoint is structured this way: FileName Cy3 Cy5 1 mu1 wt1 2 wt1 mu1 3 mu2 wt2 4 wt2 mu2 (1 and 2 are to distinguish the 2 bioreps) I'm interested not only in the DE genes between wt and mutant at each timepoint, but I would like to extract information independently from the wt and mutant time-courses, even if I don't have direct comparison between successive timepoints. Is this possible? I think there is an example in limma's user guide (chapter 16, time-course experiments), but it is for one-color array (at least, I guess), and I don't really know how to adapt the script to my case. Can anyone point me in the right direction? (please forgive my lack of statistical knowledge!) Thank you. Federico

Hi all ! I posted a message several days ago but I didn't get any answers. So I assume that the time effect in a direct comparison exp. design (across multiple timepoints) is non-estimable. Is there anyone able to confirm my assumption? I paste my old message again here: "my experiment is a 2-color direct comparison wt vs mutant, but repeated on 5 successive timepoints. each timepoint is structured this way: FileName Cy3 Cy5 1 mu1 wt1 2 wt1 mu1 3 mu2 wt2 4 wt2 mu2 (1 and 2 are to distinguish the 2 bioreps) I'm interested not only in the DE genes between wt and mutant at each timepoint, but I would like to extract information independently from the wt and mutant time-courses, even if I don't have direct comparison between successive timepoints. Is this possible? I think there is an example in limma's user guide (chapter 16, time-course experiments), but it is for one-color array (at least, I guess), and I don't really know how to adapt the script to my case." Any comment or suggestion is really appreciated. Thank you. Federico

> Date: Tue, 22 Feb 2005 17:01:42 -0800 > From: Federico Scossa <fscossa@pw.usda.gov> > Subject: [BioC] LIMMA - timecourse and direct comparison exp. design > To: bioconductor@stat.math.ethz.ch > > Hi all ! > I posted a message several days ago but I didn't get any answers. So I > assume that the time effect in a direct comparison exp. design (across > multiple timepoints) is non-estimable. > Is there anyone able to confirm my assumption? The time effect is not estimable using log-ratio analyses. You can only estimate the time effect using a single channel analysis, for example using the function lmscFit() in limma. A section on single channel analysis was added to the limma User's Guide in version 1.8.19, available from http://bioinf.wehi.edu.au/limma Gordon > I paste my old message again here: > > "my experiment is a 2-color direct comparison wt vs mutant, but repeated on > 5 successive timepoints. > each timepoint is structured this way: > > FileName Cy3 Cy5 > 1 mu1 wt1 > 2 wt1 mu1 > 3 mu2 wt2 > 4 wt2 mu2 > > (1 and 2 are to distinguish the 2 bioreps) > > I'm interested not only in the DE genes between wt and mutant at each > timepoint, but I would like to extract information independently from the > wt and mutant time-courses, even if I don't have direct comparison between > successive timepoints. Is this possible? I think there is an example in > limma's user guide (chapter 16, time-course experiments), but it is for > one-color array (at least, I guess), and I don't really know how to adapt > the script to my case." > > Any comment or suggestion is really appreciated. > Thank you. > Federico