Creating Heatmaps Of Diffrentially Expressed Genes In Agilent Two-Color Array Analysis

targets<-readTargets("HxMtargets.txt") RG <- read.maimages(targets,source="agilent",columns= list( R = "rMeanSignal", G = "gMeanSignal", Rb = "rBGUsed", Gb = "gBGUsed", Rb.real = "rBGMeanSignal", Gb.real = "gBGMeanSignal"), annotation = c("FeatureNum","Row","Col","ProbeName","ControlType","GeneName", "Description","SystematicName")) MA<-normalizeWithinArrays(RG,method="loess") f<-factor(targets$Cy3,levels=c("Control","Twowks","Tumor")) design<-model.matrix(~0+f) colnames(design) <- c("Control","Twowks","Tumor") fit<-lmFit(MA, design) contrast.matrix<-makeContrasts(Control,Twowks,Tumor,Tumor-Control,Tumor-Twowks, Twowks-Control, levels=design) fit2<-contrasts.fit(fit,contrast.matrix) fit2<-eBayes(fit2) C.top<-as.numeric(rownames(topTable(fit2, coef=1,n=50))) heatmap(MA$M[C.top,])

9.3 years ago by

Sean Davis ♦ 26k

National Institutes of Health, Bethesda, MD

Sean Davis ♦ 26k wrote:

Take a look at the gplots package, for example, at the heatmap.2 function. Something like this should give you an idea:

d = matrix(rnorm(26*26),ncol=26)
heatmap.2(d,labRow=letters[1:26],labCol=letters[1:26])

You'll just substitute your MA$M[C.top,], labRow=genenames[C.top], and labCol=samplenames. heatmap.2 has many tweaks for better visualization, so you may need to play a bit to get things just right.

ADD COMMENT • link written 9.3 years ago by Sean Davis ♦ 26k

Thanks you so much Sean! I did try this with some modifications of my own and it worked perfectly! Again, thank you.

ADD REPLY • link written 9.3 years ago by Ly • 130

Hi Ly, if you like the answer upvote and accept it.

ADD REPLY • link written 9.3 years ago by Istvan Albert ♦♦ 86k

Hi Sean,

Like the above question even I'm facing the same problem to get heatmap of differential expressed genes. Could you please hep me. I'm working on Human Gene 2.0 ST array. I have used "xps" package for the analysis.

scheme.exon <- import.exon.scheme("Scheme",filedir=scmdir,layoutfile=paste(libdir,"HuGene-2_0-st.clf",sep="/"),schemefile=paste(libdir,"HuGene-2_0-st.pgf",sep="/"),probeset=paste(anndir,"HuGene-2_0-st-v1.na35.hg19.probeset.csv",sep="/"),transcript=paste(anndir,"HuGene-2_0-st-v1.na35.hg19.transcript.csv",sep="/"),verbose=TRUE)

##Reading CEL Info
#Import CEL-files by giving the path where all .CEL files are present

celfiles <- dir(path = "D:/GeneST_Analyse/ProjectName/CELfiles", pattern = "*.CEL$", all.files = FALSE, full.names = FALSE, recursive = FALSE, ignore.case = FALSE)
data.exon <- import.data(scheme.exon,"tmp_data_exon",filedir=rootdir,celfiles=celfiles,celdir=celdir,verbose=FALSE)

unlist(treeNames(data.exon))

[1] "X_8__HuGene_2_0_st_.cel" "E__HuGene_2_0_st_.cel"  
[3] "GG7__HuGene_2_0_st_.cel" "J__HuGene_2_0_st_.cel"  
[5] "O__HuGene_2_0_st_.cel"   "T__HuGene_2_0_st_.cel"

data.rma <- rma(data.exon, "tmp_exonRMA", filedir=rootdir, verbose=FALSE, exonlevel="affx+core", option="transcript", background="antigenomic")

>data.rma

UNIT_ID   UN       X       E       GG7     J     O
0      17127159 353.568 335.295 221.717 815.654 684.85
1      17127161 11.0842 7.01459 7.33511 11.2121 12.6268
2      17127163 221.647 226.774 136.274 431.32  392.533
3      17127165 5.02182 3.41172 4.12834 6.90306 4.91183

unifltr <- UniFilter(foldchange=c(1.5,"both"), unifilter=c(0.05,"pval"))

rma.ufr <- unifilter(data.rma, "tmp_exonUnifilter", filedir=rootdir , filter=unifltr, group=c("GrpA","GrpA","GrpA","GrpB","GrpB","GrpB"), xps.fltr=rma.pfr, verbose=FALSE)

tmp2 <- validData(rma.ufr)

tmp2 has differential expressed genes.

UN          ID    St      M1    M2       SE    DOF  PV        PA            FC
17127159    0   -5.9    297.3   765.7   0.22    4   0.003   0.00389231  2.57536
17127163    2   -3.87   189.914 492.307 0.3548  4   0.0179  0.01795     2.59226
17127167    4   -3.8908 339.136 855.276 0.3429  4   0.0176  0.017       2.52192
17127171    6   -3.922  390.44  986.365 0.340   4   0.0172179   0.01721 2.52627
17127175    8   -4.715  536.072 1210.65 0.2492  4   0.00920158  0.00920 2.258

How can I get a heatmap for this?

ADD REPLY • link modified 4.9 years ago by Istvan Albert ♦♦ 86k • written 4.9 years ago by Vasu • 530

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