
Jim, 
     I need to do an analysis of expression differential of genes. I do not know if using M values ​​or log-ratios.

Regards,


> Date: Mon, 19 Sep 2011 09:06:03 -0400
> From: jmacdon@med.umich.edu
> To: verotapia@alumnos.utalca.cl
> CC: yong.li@zbsa.uni-freiburg.de; bioconductor@r-project.org
> Subject: Re: [BioC] process one color microarray
> 
> Hi Paz,
> 
> On 9/18/2011 11:20 AM, Paz Tapia Ramirez wrote:
> > Hi JIM.
> > When you say"trt1" "trt2" you mean the column GErep in the targets file ?
> 
> I don't know anything about your targets file, as you didn't show us 
> what that looks like.
> 
> And as Yong noted, even though you claim a one-color array, you appear 
> to be doing something other than a conventional one-color analysis. So 
> first you need to clarify exactly what you are trying to do with this 
> analysis, what kind of data these are, what your targets file looks 
> like, etc.
> 
> Best,
> 
> Jim
> 
> >
> >  > Date: Fri, 16 Sep 2011 13:32:30 +0200
> >  > From: yong.li@zbsa.uni-freiburg.de
> >  > To: jmacdon@med.umich.edu
> >  > CC: verotapia@alumnos.utalca.cl; bioconductor@r-project.org
> >  > Subject: Re: [BioC] process one color microarray
> >  >
> >  > Hi Paz, hi Jim,
> >  >
> >  > I think there is a problem here. Jim, you have given detailed
> >  > explanation of making design matrix and do various comparisons, but they
> >  > are based on the assumption that the input given to lmFit is log
> >  > expression values from one color array. The problem is that, based on
> >  > Paz's code, in the MAList MA1 the M values are log 2 ratios of mean to
> >  > processed signals of the green channel (of course background corrected
> >  > and quantile normalized). I doubt very much that that piece of codes
> >  > does what Paz wants to do. However, it's also possible that I missed
> >  > something.
> >  >
> >  > Best regards,
> >  > Yong
> >  >
> >  > James W. MacDonald wrote:
> >  > > Hi Paz,
> >  > >
> >  > > On 9/14/2011 5:55 PM, Paz Tapia Ramirez wrote:
> >  > >> Hello, I have a question. I'm Working with one-color microarrays . I
> >  > >> worked in 4 conditions different and each condition I have 4
> >  > >> replicates. Now, my question is when I load the files to
> >  > >> Bioconductor, I load as follows:
> >  > >> my.filenames<- c ("Condic1_repl1.txt",
> >  > >> "Condic1_repl2.txt",
> >  > >> "Condic1_repl3.txt",
> >  > >> "Condic1_repl4.txt",
> >  > >> "Condic2_repl1.txt",
> >  > >> "Condic2_repl2.txt",
> >  > >> "Condic2_repl3.txt",
> >  > >> "Condic2_repl4.txt",
> >  > >> "Condic3_repl1.txt"
> >  > >> "Condic3_repl2.txt",
> >  > >> "Condic3_repl3.txt",
> >  > >> "Condic3_repl4.txt",
> >  > >> "Condic3_repl1.txt",
> >  > >> "Condic3_repl2.txt",
> >  > >> "Condic3_repl3.txt",
> >  > >> "Condic3_repl4.txt")
> >  > >>
> >  > >> Subsequently, I realize the normalization procedure and some
> >  > >> statistical calculations:
> >  > >> one.col1<-list (R = "gMeanSignal" G = "gProcessedSignal"
> >  > >> Rb = "gBGMedianSignal", Gb = "gProcessedBackground")
> >  > >> RG1<- read.maimages (my.filenames, source = "agilent", columns =
> >  > >> one.col1, dec =".")
> >  > >> RG1<- backgroundCorrect (RG1, method = "half", offset = 50)
> >  > >> MA1<- normalizeBetweenArrays (RG1, method = "quantile")
> >  > >> fit1<- lmFit (MA1, design = NULL)
> >  > >
> >  > > The design matrix indicates to lmFit() what is control and what is
> >  > > treatment. When you specify a NULL design matrix, lmFit() will just
> > use
> >  > > a vector of 1s, which would be fine if you had two-color chips and no
> >  > > dye-swaps. Otherwise, you are just testing the hypothesis that the
> >  > > average expression of all samples is not equal to zero (which
> > obviously
> >  > > isn't correct).
> >  > >
> >  > > So if you have four conditions with four replicates (and I am assuming
> >  > > here that they are Biological replicates, not just different
> > aliquots of
> >  > > the same sample), you want a design matrix with four columns. The
> >  > > simplest such design matrix (to me, anyway), computes the mean
> >  > > expression for each group, and then you can just make the comparisons
> >  > > you want.
> >  > >
> >  > > cond <- factor(rep(1:4, each = 4))
> >  > > design <- model.matrix(~ 0 + cond)
> >  > > colnames(design) <- c("trt1","trt2","trt3","trt4")
> >  > >
> >  > > contrast <- makeContrasts(trt2-trt1, trt3-trt1, trt4-trt1, trt3-trt2,
> >  > > trt4-trt2, trt4-trt3, levels = design)
> >  > > fit <- lmFit(Ma1, design)
> >  > > fit1 <- contrasts.fit(fit, contrast)
> >  > > fit1 <- eBayes(fit1)
> >  > >
> >  > > which will make all possible comparisons.
> >  > >
> >  > > Alternatively, if you just want to compare all treatments to a control
> >  > > (and assuming your control is trt1).
> >  > >
> >  > > design <- model.matrix(~cond)
> >  > > fit <- lmFit(Ma1, design)
> >  > > fit1 <- eBayes(fit)
> >  > >
> >  > > In this case, all coefficients in the model will be e.g., trt2-trt1,
> >  > > trt3-trt1, trt4-trt1, so you don't have to specify contrasts directly.
> >  > >
> >  > > Best,
> >  > >
> >  > > Jim
> >  > >
> >  > >
> >  > >
> >  > >> fit1<- eBay (fit1)
> >  > >>
> >  > >> But my question is: how I can specify to bioconductor which files
> >  > >> correspond to Control, or which correspond to microarrays with
> >  > >> treatment?
> >  > >>
> >  > >> Regards, Paz
> >  > >>
> >  > >> [[alternative HTML version deleted]]
> >  > >>
> >  > >> _______________________________________________
> >  > >> Bioconductor mailing list
> >  > >> Bioconductor@r-project.org
> >  > >> https://stat.ethz.ch/mailman/listinfo/bioconductor
> >  > >> Search the archives:
> >  > >> http://news.gmane.org/gmane.science.biology.informatics.conductor
> >  > >
> 
> -- 
> James W. MacDonald, M.S.
> Biostatistician
> Douglas Lab
> University of Michigan
> Department of Human Genetics
> 5912 Buhl
> 1241 E. Catherine St.
> Ann Arbor MI 48109-5618
> 734-615-7826
> **********************************************************
> Electronic Mail is not secure, may not be read every day, and should not be used for urgent or sensitive issues 
> 
 		 	   		  
	[[alternative HTML version deleted]]

