Hello All, I am trying to fit the data to the linear model to get the differential expressed gene list. The experiment has a loop design with 12 hybridizations consisting of 6 samples (each with 4 biological replicates). After I run the following code, i encountered the warning message from "eBayes" function. Then when i run the topTable function, the resulting table contains duplicated entries as shown below. I also enclosed the "targets", "design" and the MA object "MA.AB" for your information. Can someone help me idntify the problem? Thanks a lot! Jia >design.names=list(c("slide1","slide2","slide3","slide4","slide5","sl ide6","slide7","slide8","slide9","slide10","slide11","slide12"),c("DB- DM","DM-NM","DM-DT","NM-NB","NT-NM")) >design=matrix(c(1,1,0,1,0,1,0,0,0,0,0,1,0,0,0,0,0,1,0,0,0,1,-1,0,-1, -1,0,-1,0,0,-1,-1,0,-1,0,0,0,0,-1,-1,0,-1,0,0,0,0,0,0,1,0,0,-1,1,0,1,0 ,0,0,0,1),12,5,byrow=T,dimnames=design.names) > fit<-lmFit(MA.AB,design=design,weights=MA.AB$weights, p.adjust="Hochberg") > fit<-eBayes(fit) *Warning message: Estimation of var.prior failed - set to default value in: ebayes(fit = fit, proportion = proportion, stdev.coef.lim = stdev.coef.lim)* > topTable(fit[,1]) Block Row Column ID Name Status 4080 7 21 7 MZ00043373 TC220219 gene 32550 7 21 7 MZ00043373 TC220219 gene 13125 22 24 5 MZ00028432 TC225126 gene 41595 22 24 5 MZ00028432 TC225126 gene 23889 40 22 16 MZ00027664 TC225103 gene 52359 40 22 16 MZ00027664 TC225103 gene 15428 26 20 1 MZ00027244 TC239087 gene 43898 26 20 1 MZ00027244 TC239087 gene 27756 47 21 8 MZ00043374 TC206176 gene 56226 47 21 8 MZ00043374 TC206176 gene Annotation 4080 cellulose synthase {Oryza sativa (japonica cultivar-group);} 32550 cellulose synthase {Oryza sativa (japonica cultivar-group);} 13125 hypothetical protein {Oryza sativa (japonica cultivar-group);} ^|^GB|BAB92324.1|20804634|AP003253 hypothetical protein {Oryza sativa (japonica cultivar-group);} 41595 hypothetical protein {Oryza sativa (japonica cultivar-group);} ^|^GB|BAB92324.1|20804634|AP003253 hypothetical protein {Oryza sativa (japonica cultivar-group);} 23889 putative CER1 {Oryza sativa (japonica cultivar-group);} ^|^GB|AAG21908.1|10716610|AC026815 putative CER1 {Oryza sativa (japonica cultivar-group);} 52359 putative CER1 {Oryza sativa (japonica cultivar-group);} ^|^GB|AAG21908.1|10716610|AC026815 putative CER1 {Oryza sativa (japonica cultivar-group);} 15428 OSJNBa0029H02.22 {Oryza sativa (japonica cultivar- group);} ^|^GB|CAD41488.1|21740509|OSJN00025 OSJNBa0029H02.27 {Oryza sativa (japonica cultivar-group);} 43898 OSJNBa0029H02.22 {Oryza sativa (japonica cultivar- group);} ^|^GB|CAD41488.1|21740509|OSJN00025 OSJNBa0029H02.27 {Oryza sativa (japonica cultivar-group);} 27756 cellulose synthase-6 {Zea mays;} 56226 cellulose synthase-6 {Zea mays;} logFC AveExpr t P.Value adj.P.Val B 4080 -2.446885 9.317316 -8.446857 2.037268e-05 0.2538640 2.079626317 32550 -2.446885 9.317316 -8.446857 2.037268e-05 0.2538640 2.079626317 13125 1.913508 11.108700 5.139045 2.808850e-04 0.5331159 0.539694800 41595 1.913508 11.108700 5.139045 2.808850e-04 0.5331159 0.539694800 23889 -1.624284 11.770228 -5.012985 3.443437e-04 0.5331159 0.372559874 52359 -1.624284 11.770228 -5.012985 3.443437e-04 0.5331159 0.372559874 15428 -1.456442 9.134642 -5.143359 5.125077e-04 0.5331159 -0.006356757 43898 -1.456442 9.134642 -5.143359 5.125077e-04 0.5331159 -0.006356757 27756 -2.108918 8.578181 -6.546458 4.367518e-04 0.5331159 -0.091344144 56226 -2.108918 8.578181 -6.546458 4.367518e-04 0.5331159 -0.091344144 > targets SlideNumber FileName Cy3 Cy5 1 1 A160.gpr DB NB 2 2 A164.gpr DB DM 3 3 A165.gpr DM NM 4 4 A166.gpr DM DT 5 5 A167.gpr DT NT 6 6 A168.gpr DT DB 7 7 A169.gpr NB DB 8 8 A170.gpr NB NT 9 9 A171.gpr NM DM 10 10 A077.gpr NM NB 11 11 A078.gpr NT DT 12 12 A079.gpr NT NM > design DB-DM DM-NM DM-DT NM-NB NT-NM slide1 1 1 0 1 0 slide2 1 0 0 0 0 slide3 0 1 0 0 0 slide4 0 0 1 0 0 slide5 0 1 -1 0 -1 slide6 -1 0 -1 0 0 slide7 -1 -1 0 -1 0 slide8 0 0 0 -1 -1 slide9 0 -1 0 0 0 slide10 0 0 0 1 0 slide11 0 -1 1 0 1 slide12 0 0 0 0 1 > MA.AB An object of class "MAList" $weights B167 B168 B169 B170 B177 B180 B181 B182 B183 B070 B072 B073 [1,] 0 0 0 0 0 0 0 0 0 0 0 0 [2,] 0 0 0 0 0 0 0 0 0 0 0 0 [3,] 0 0 0 0 0 0 0 0 0 0 0 0 [4,] 0 0 0 0 0 0 0 0 0 0 0 0 [5,] 0 1 1 1 1 1 0 1 0 0 1 1 56935 more rows ... $targets SlideNumber FileName Cy3 Cy5 1 1 B167.gpr DB NB 2 2 B168.gpr DB DM 3 3 B169.gpr DM NM 4 4 B170.gpr DM DT 5 5 B177.gpr DT NT 7 more rows ... $genes Block Row Column ID Name Status 1 1 1 1 MZ00002380 AZM4_14521 gene 2 1 1 2 MZ00002386 AZM4_49988 gene 3 1 1 3 MZ00002404 AZM4_100937 gene 4 1 1 4 MZ00002410 AZM4_135577 gene 5 1 1 5 MZ00002428 AZM4_79828 gene Annotation 1 phosphatidylinositol 3,5-kinase- like {Oryza sativa (japonica cultivar-group);} 2 OSJNBa0087O24.3 {Oryza sativa (japonica cultivar-group);} 3 hypothetical protein similar to Oryza sativa chromosome 1, P0431G06.13 {Oryza sativa (japonica cultivar-group);} 4 putative phosphatase {Oryza sativa (japonica cultivar-group);} 5 putative flavonol 3-O-glucosyltransferase {Oryza sativa (japonica cultivar-group);} 56935 more rows ... $source [1] "genepix" $printer $ngrid.r [1] 12 $ngrid.c [1] 4 $nspot.r [1] 26 $nspot.c [1] 26 attr(,"class") [1] "PrintLayout" $M B167 B168 B169 B170 B177 B180 [1,] -1.3779913 -1.8833111 -1.62708741 -1.3495565 -1.1180732 -0.5628514 [2,] -0.4571145 -0.6198667 -0.06527221 -0.5730541 -0.6867249 -0.6093460 [3,] -0.2940607 -0.2293377 -0.20736120 -0.9225977 -1.0349128 -0.8617654 [4,] -1.4274691 -1.1814338 -1.45430688 -0.5903722 -0.8967976 -1.2961053 [5,] 0.9068494 0.1585617 -0.10686769 0.3406360 -0.5760433 -1.0153630 B181 B182 B183 B070 B072 B073 [1,] NA -1.8638750 -0.9362995 -2.577714 -1.99677057 -0.1696838 [2,] -1.2031207 -0.1840673 -0.9566250 -2.154500 -0.52701635 -0.2899944 [3,] 0.2947026 1.2401698 NA -1.686424 -0.01693860 -0.1007202 [4,] -1.4675358 -2.6755268 NA -2.531036 -0.69658202 -2.5460154 [5,] 1.0599882 1.4856234 1.4375392 -1.177492 0.49266749 1.4147140 56935 more rows ... $A B167 B168 B169 B170 B177 B180 B181 B182 [1,] 7.633485 6.241182 6.675137 6.402408 6.370169 6.356133 NA 5.784979 [2,] 8.937003 6.628853 5.813245 6.034256 6.138358 5.515603 5.656917 5.668056 [3,] 5.512734 6.183253 6.367349 5.099702 5.612979 5.124473 5.581835 4.329897 [4,] 7.185585 6.401859 5.634371 6.137675 6.210610 6.660527 5.355347 5.621532 [5,] 7.074069 8.971200 9.019419 9.814719 9.862116 8.833574 7.790254 8.648343 B183 B070 B072 B073 [1,] 6.905622 6.242238 5.785340 6.237580 [2,] 7.115129 6.069744 5.617608 6.084069 [3,] NA 5.322445 5.309317 5.177219 [4,] NA 6.566366 5.293450 4.902802 [5,] 8.803978 5.990159 8.700017 7.253411 56935 more rows ...