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<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>(sorry there was some missing information)</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>Hello all!</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>I am a beginner user of R and Bioconductor, sorry if my questions have already been discussed previously.</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>I am studying the effects of a new hypoglycaemic drug for the treatment of diabetes and I have done this classical study:</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-INDENT: 35.4pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>4 different groups:</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt 35.4pt; TEXT-INDENT: 35.4pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>1.- Healthy untreated</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt 35.4pt; TEXT-INDENT: 35.4pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>2.- Healthy treated</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt 35.4pt; TEXT-INDENT: 35.4pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>3.- Diabetic untreated</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt 35.4pt; TEXT-INDENT: 35.4pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>4.- Diabetic treated</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>With 3 biological replicates of each group, therefore I have done 12 arrays (Affymetrix).</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>I have treated the raw data with the package RMA of Bioconductor according to the article ”Exploration, normalization and summaries of high density oligonucleotide array probe level data” (Background=RMA, Normalization=quantiles, PM=PMonly, Summarization=medianpolish).</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>I am currently trying to analyse the object eset with the package LIMMA of Bioconductor. I want to know what genes are differentially expressed due to diabetes,<SPAN style="mso-spacerun: yes"> </SPAN>to the treatment and to the combination of both (diabetes + treatment), being therefore an statistic analysis similar to a two-ways ANOVA).</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>So, my questions are:</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>1.- I have created a PhenoData in RMA, will the covariates of the PhenoData have any influence in the analysis of LIMMA?</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>2.- Are these commands correct to get these results? (see below) In the command TopTable, the output of coef=1 are the genes characteristics of diabetes? </FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>3.- If I do not see any effect of the treatment in the healthy untreated rats should I design the matrix differently? Something similar to a one-way-ANOVA, considering differently the four groups:</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>( > design<-model.matrix(~ -1+factor(c(1,1,1,2,2,2,3,3,3,4,4,4)))<SPAN style="mso-spacerun: yes"> </SPAN>).</FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>5.- Any other idea?</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>Thank you very much for your time and your suggestions.</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>Yours sincerely,</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT size=3><FONT face="Times New Roman"> <?xml:namespace prefix = o ns = "urn:schemas-microsoft-com:office:office" /><o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>Jordi Altirriba (PhD student, Hospital Clínic – IDIBAPS, Barcelona, Spain)</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT size=3><FONT face="Times New Roman"> <o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>> design<-cbind("disease"=c(1,1,1,1,1,1,0,0,0,0,0,0),"treatment"=c(0,0,0,1,1,1,0,0,0,1,1,1))</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN style="mso-ansi-language: ES"><FONT size=3><FONT face="Times New Roman">> fit<-lmFit(eset,design)<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman">> contrast.matrix<-cbind("diabetes"=c(1,0),"drug"=c(0,1),"diabetes-drug"=c(1,1))<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN style="mso-ansi-language: ES"><FONT size=3><FONT face="Times New Roman">> rownames(contrast.matrix)<-colnames(design)<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN style="mso-ansi-language: ES"><FONT size=3><FONT face="Times New Roman">> design<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN style="mso-ansi-language: ES"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>disease treatment<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN style="mso-ansi-language: ES"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>[1,]<SPAN style="mso-spacerun: yes"> </SPAN>1<SPAN style="mso-spacerun: yes"> </SPAN>0<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-ansi-language: ES"><SPAN style="mso-spacerun: yes"> </SPAN></SPAN><SPAN lang=FR style="mso-ansi-language: FR">[2,]<SPAN style="mso-spacerun: yes"> </SPAN>1<SPAN style="mso-spacerun: yes"> </SPAN>0<o:p></o:p></SPAN></FONT></FONT></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>[3,]<SPAN style="mso-spacerun: yes"> </SPAN>1<SPAN style="mso-spacerun: yes"> </SPAN>0<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>[4,]<SPAN style="mso-spacerun: yes"> </SPAN>1<SPAN style="mso-spacerun: yes"> </SPAN>1<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>[5,]<SPAN style="mso-spacerun: yes"> </SPAN><SPAN style="mso-spacerun: yes"> </SPAN>1<SPAN style="mso-spacerun: yes"> </SPAN>1<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>[6,]<SPAN style="mso-spacerun: yes"> </SPAN>1<SPAN style="mso-spacerun: yes"> </SPAN>1<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>[7,]<SPAN style="mso-spacerun: yes"> </SPAN>0<SPAN style="mso-spacerun: yes"> </SPAN>0<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>[8,]<SPAN style="mso-spacerun: yes"> </SPAN>0<SPAN style="mso-spacerun: yes"> </SPAN>0<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>[9,]<SPAN style="mso-spacerun: yes"> </SPAN>0<SPAN style="mso-spacerun: yes"> </SPAN>0<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman">[10,]<SPAN style="mso-spacerun: yes"> </SPAN>0<SPAN style="mso-spacerun: yes"> </SPAN>1<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman">[11,]<SPAN style="mso-spacerun: yes"> </SPAN>0<SPAN style="mso-spacerun: yes"> </SPAN>1<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman">[12,]<SPAN style="mso-spacerun: yes"> </SPAN>0<SPAN style="mso-spacerun: yes"> </SPAN>1<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman">> contrast.matrix<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>diabetes drug diabetes-drug<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman">diabetes<SPAN style="mso-spacerun: yes"> </SPAN>1<SPAN style="mso-spacerun: yes"> </SPAN>0<SPAN style="mso-spacerun: yes"> </SPAN>1<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman">tratamiento<SPAN style="mso-spacerun: yes"> </SPAN>0<SPAN style="mso-spacerun: yes"> </SPAN>1<SPAN style="mso-spacerun: yes"> </SPAN>1<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=FR style="mso-ansi-language: FR"><FONT size=3><FONT face="Times New Roman">> fit2<-contrasts.fit(fit,contrast.matrix)<o:p></o:p></FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>> fit2<-eBayes(fit2)</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>> clas<-classifyTests(fit2)</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>> vennDiagram(clas)</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>> topTable(fit2,number=100,genelist=geneNames(eset),coef=1,adjust="fdr")</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT size=3><FONT face="Times New Roman"><SPAN style="mso-spacerun: yes"> </SPAN>Name<SPAN style="mso-spacerun: yes"> </SPAN>M<SPAN style="mso-spacerun: yes"> </SPAN>t<SPAN style="mso-spacerun: yes"> </SPAN>P.Value<SPAN style="mso-spacerun: yes"> </SPAN>B</FONT></FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>11590<SPAN style="mso-spacerun: yes"> </SPAN>1387471_at 19.32575 9.442926 2.743122e-17 29.95299</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>2500<SPAN style="mso-spacerun: yes"> </SPAN>1369951_at 19.24748 9.404683 2.743122e-17 29.66737</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>10652<SPAN style="mso-spacerun: yes"> </SPAN>1384778_at 19.22968 9.395981 2.743122e-17 29.60254</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>…</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>> sink("limma-diabetes.txt")</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>> topTable(fit2,number=1000,genelist=geneNames(eset),coef=1,adjust="fdr")</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT face="Times New Roman" size=3>> sink()</FONT></SPAN></P>
<P class=MsoNormal style="MARGIN: 0cm 0cm 0pt; TEXT-ALIGN: justify"><SPAN lang=EN-GB><FONT size=3><FONT face="Times New Roman"> <o:p></o:p></FONT></FONT></SPAN></P></DIV></div><br clear=all><hr>¿Todavía no conoces el nuevo MSN Messenger? <a href="http://g.msn.com/8HMBESES/2728??PS=">Descárgatelo gratis aquí</a> </html>