HELP! lmFit and duplicateCorrelation
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Tiandao Li ▴ 260
@tiandao-li-2372
Last seen 10.2 years ago
Hello, I had arrays with 4 replicate spots per gene. I used limma package for data analysis. > targets SlideNumber FileName Cy3 Cy5 Name Field1 1 13617731 WBM WC Field1 Field2 2 13617730 WBM WC Field2 Field3 3 13617724 WC WBM Field3 Field4 4 13617627 WC WBM Field4 Field5 5 13617626 WBM WC Field5 After read in data, normalization, I used the following codes for within-array replicate spots. design <- modelMatrix(targets, ref="WC") corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow computation! fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, method="ls") fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, method="robust") # eBayes fit5 <- eBayes(fit) fit6 <- eBayes(fit2) Then, topTable(fit5, number=30, adjust="BH") gives me a list of differentially expressed genes. However, some of genes show up mutiple times, such as 37A-C02.g in the list. According to limma guide, 11.6, gene names should appear only once. Block Row Column ID Name 597 4 15 15 37B-B10.g P25782 519 4 2 17 37A-C02.g P00765 531 4 4 17 37A-C02.g P00765 314 2 24 18 37A-B08.g AC186398 308 2 23 19 35B-F12.g AC122261 513 4 1 17 37A-C02.g P00765 Another question, is there any difference between differentially expressed genes using methods of "ls" and "robust" in lmFit? > sessionInfo() R version 2.6.0 (2007-10-03) i486-pc-linux-gnu locale: LC_CTYPE=en_US.UTF-8;LC_NUMERIC=C;LC_TIME=en_US.UTF-8;LC_COLLATE=en_US .UTF-8;LC_MONETARY=en_US.UTF-8;LC_MESSAGES=en_US.UTF-8;LC_PAPER=en_US. UTF-8;LC_NAME=C;LC_ADDRESS=C;LC_TELEPHONE=C;LC_MEASUREMENT=en_US.UTF-8 ;LC_IDENTIFICATION=C attached base packages: [1] stats graphics grDevices utils datasets methods base other attached packages: [1] MASS_7.2-37 statmod_1.3.1 limma_2.12.0 loaded via a namespace (and not attached): [1] rcompgen_0.1-15 Best wishes, Tiandao
Normalization limma Normalization limma • 2.6k views
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Ido M. Tamir ▴ 320
@ido-m-tamir-1268
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On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > Hello, > > I had arrays with 4 replicate spots per gene. I used limma package for > data analysis. > > > targets > > SlideNumber FileName Cy3 Cy5 Name > Field1 1 13617731 WBM WC Field1 > Field2 2 13617730 WBM WC Field2 > Field3 3 13617724 WC WBM Field3 > Field4 4 13617627 WC WBM Field4 > Field5 5 13617626 WBM WC Field5 > > After read in data, normalization, I used the following codes for > within-array replicate spots. > > design <- modelMatrix(targets, ref="WC") > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow computation! > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > method="ls") > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > method="robust") Dear Tiandao, unless the replicate spots are consecutive you have to give a spacing argument to indicate what the replicate spots actually are. If they are randomly spotted on the array, you would have to rearrange them somehow and then use the appropriate spacing. 519 4 2 17 37A-C02.g P00765 531 4 4 17 37A-C02.g P00765 doesn't look as if the replicate spots are one after the other. HTH best wishes, ido
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Hello Ido, Block Row Column ID Name 519 4 2 17 37A-C02.g P00765 531 4 4 17 37A-C02.g P00765 513 4 1 17 37A-C02.g P00765 For 37A-C02.g, I only selected the first couple of genes from the topTable list, all 4 replicate spots are printed consecutively in the same column. Thanks for your reply. Tiandao On Tue, 9 Oct 2007, Ido M. Tamir wrote: On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > Hello, > > I had arrays with 4 replicate spots per gene. I used limma package for > data analysis. > > > targets > > SlideNumber FileName Cy3 Cy5 Name > Field1 1 13617731 WBM WC Field1 > Field2 2 13617730 WBM WC Field2 > Field3 3 13617724 WC WBM Field3 > Field4 4 13617627 WC WBM Field4 > Field5 5 13617626 WBM WC Field5 > > After read in data, normalization, I used the following codes for > within-array replicate spots. > > design <- modelMatrix(targets, ref="WC") > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow computation! > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > method="ls") > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > method="robust") Dear Tiandao, unless the replicate spots are consecutive you have to give a spacing argument to indicate what the replicate spots actually are. If they are randomly spotted on the array, you would have to rearrange them somehow and then use the appropriate spacing. 519 4 2 17 37A-C02.g P00765 531 4 4 17 37A-C02.g P00765 doesn't look as if the replicate spots are one after the other. HTH best wishes, ido _______________________________________________ Bioconductor mailing list Bioconductor at stat.math.ethz.ch https://stat.ethz.ch/mailman/listinfo/bioconductor Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor
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At 09:19 AM 10/9/2007, Tiandao Li wrote: >Hello Ido, > > Block Row Column ID Name >519 4 2 17 37A-C02.g P00765 >531 4 4 17 37A-C02.g P00765 >513 4 1 17 37A-C02.g P00765 > >For 37A-C02.g, I only selected the first couple of genes from the topTable >list, all 4 replicate spots are printed consecutively in the same column. This is not the same as consecutively spacing in ROWS, which is what the default value of spacing=1 means. Your spacing is probably the number of spots per row on the array. Check MA$genes to see how many rows of the data matrix separate your duplicates. Jenny >Thanks for your reply. > >Tiandao > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > Hello, > > > > I had arrays with 4 replicate spots per gene. I used limma package for > > data analysis. > > > > > targets > > > > SlideNumber FileName Cy3 Cy5 Name > > Field1 1 13617731 WBM WC Field1 > > Field2 2 13617730 WBM WC Field2 > > Field3 3 13617724 WC WBM Field3 > > Field4 4 13617627 WC WBM Field4 > > Field5 5 13617626 WBM WC Field5 > > > > After read in data, normalization, I used the following codes for > > within-array replicate spots. > > > > design <- modelMatrix(targets, ref="WC") > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow computation! > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > method="ls") > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > method="robust") > >Dear Tiandao, > >unless the replicate spots are consecutive you have to give a spacing >argument to indicate what the replicate spots actually are. >If they are randomly spotted on the array, you would have to rearrange >them somehow and then use the appropriate spacing. > >519 4 2 17 37A-C02.g P00765 >531 4 4 17 37A-C02.g P00765 > >doesn't look as if the replicate spots are one after the other. > >HTH best wishes, >ido > >_______________________________________________ >Bioconductor mailing list >Bioconductor at stat.math.ethz.ch >https://stat.ethz.ch/mailman/listinfo/bioconductor >Search the archives: >http://news.gmane.org/gmane.science.biology.informatics.conductor > >_______________________________________________ >Bioconductor mailing list >Bioconductor at stat.math.ethz.ch >https://stat.ethz.ch/mailman/listinfo/bioconductor >Search the archives: >http://news.gmane.org/gmane.science.biology.informatics.conductor
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Dear Jenny, I checked the MA$genes, the following is part of the list 1 45Rev2-A03.g .... 25 45Rev2-A03.g ... 49 45Rev2-A03.g 23 rows between 2 replicate spots, so spacing=24? Thansk, Tiandao On Tue, 9 Oct 2007, Jenny Drnevich wrote: At 09:19 AM 10/9/2007, Tiandao Li wrote: >Hello Ido, > > Block Row Column ID Name >519 4 2 17 37A-C02.g P00765 >531 4 4 17 37A-C02.g P00765 >513 4 1 17 37A-C02.g P00765 > >For 37A-C02.g, I only selected the first couple of genes from the topTable >list, all 4 replicate spots are printed consecutively in the same column. This is not the same as consecutively spacing in ROWS, which is what the default value of spacing=1 means. Your spacing is probably the number of spots per row on the array. Check MA$genes to see how many rows of the data matrix separate your duplicates. Jenny >Thanks for your reply. > >Tiandao > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > Hello, > > > > I had arrays with 4 replicate spots per gene. I used limma package for > > data analysis. > > > > > targets > > > > SlideNumber FileName Cy3 Cy5 Name > > Field1 1 13617731 WBM WC Field1 > > Field2 2 13617730 WBM WC Field2 > > Field3 3 13617724 WC WBM Field3 > > Field4 4 13617627 WC WBM Field4 > > Field5 5 13617626 WBM WC Field5 > > > > After read in data, normalization, I used the following codes for > > within-array replicate spots. > > > > design <- modelMatrix(targets, ref="WC") > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow computation! > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > method="ls") > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > method="robust") > >Dear Tiandao, > >unless the replicate spots are consecutive you have to give a spacing >argument to indicate what the replicate spots actually are. >If they are randomly spotted on the array, you would have to rearrange >them somehow and then use the appropriate spacing. > >519 4 2 17 37A-C02.g P00765 >531 4 4 17 37A-C02.g P00765 > >doesn't look as if the replicate spots are one after the other. > >HTH best wishes, >ido > >_______________________________________________ >Bioconductor mailing list >Bioconductor at stat.math.ethz.ch >https://stat.ethz.ch/mailman/listinfo/bioconductor >Search the archives: >http://news.gmane.org/gmane.science.biology.informatics.conductor > >_______________________________________________ >Bioconductor mailing list >Bioconductor at stat.math.ethz.ch >https://stat.ethz.ch/mailman/listinfo/bioconductor >Search the archives: >http://news.gmane.org/gmane.science.biology.informatics.conductor _______________________________________________ Bioconductor mailing list Bioconductor at stat.math.ethz.ch https://stat.ethz.ch/mailman/listinfo/bioconductor Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor
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At 10:38 AM 10/9/2007, Tiandao Li wrote: >Dear Jenny, > >I checked the MA$genes, the following is part of the list > >1 45Rev2-A03.g >.... >25 45Rev2-A03.g >... >49 45Rev2-A03.g > >23 rows between 2 replicate spots, so spacing=24? Yes. >Thansk, > >Tiandao > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > >At 09:19 AM 10/9/2007, Tiandao Li wrote: > >Hello Ido, > > > > Block Row Column ID Name > >519 4 2 17 37A-C02.g P00765 > >531 4 4 17 37A-C02.g P00765 > >513 4 1 17 37A-C02.g P00765 > > > >For 37A-C02.g, I only selected the first couple of genes from the topTable > >list, all 4 replicate spots are printed consecutively in the same column. > >This is not the same as consecutively spacing in ROWS, which is what >the default value of spacing=1 means. Your spacing is probably the >number of spots per row on the array. Check MA$genes to see how many >rows of the data matrix separate your duplicates. > >Jenny > > > > >Thanks for your reply. > > > >Tiandao > > > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > > > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > > Hello, > > > > > > I had arrays with 4 replicate spots per gene. I used limma package for > > > data analysis. > > > > > > > targets > > > > > > SlideNumber FileName Cy3 Cy5 Name > > > Field1 1 13617731 WBM WC Field1 > > > Field2 2 13617730 WBM WC Field2 > > > Field3 3 13617724 WC WBM Field3 > > > Field4 4 13617627 WC WBM Field4 > > > Field5 5 13617626 WBM WC Field5 > > > > > > After read in data, normalization, I used the following codes for > > > within-array replicate spots. > > > > > > design <- modelMatrix(targets, ref="WC") > > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow computation! > > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > method="ls") > > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > method="robust") > > > >Dear Tiandao, > > > >unless the replicate spots are consecutive you have to give a spacing > >argument to indicate what the replicate spots actually are. > >If they are randomly spotted on the array, you would have to rearrange > >them somehow and then use the appropriate spacing. > > > >519 4 2 17 37A-C02.g P00765 > >531 4 4 17 37A-C02.g P00765 > > > >doesn't look as if the replicate spots are one after the other. > > > >HTH best wishes, > >ido > > > >_______________________________________________ > >Bioconductor mailing list > >Bioconductor at stat.math.ethz.ch > >https://stat.ethz.ch/mailman/listinfo/bioconductor > >Search the archives: > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > >_______________________________________________ > >Bioconductor mailing list > >Bioconductor at stat.math.ethz.ch > >https://stat.ethz.ch/mailman/listinfo/bioconductor > >Search the archives: > >http://news.gmane.org/gmane.science.biology.informatics.conductor > >_______________________________________________ >Bioconductor mailing list >Bioconductor at stat.math.ethz.ch >https://stat.ethz.ch/mailman/listinfo/bioconductor >Search the archives: >http://news.gmane.org/gmane.science.biology.informatics.conductor Jenny Drnevich, Ph.D. Functional Genomics Bioinformatics Specialist W.M. Keck Center for Comparative and Functional Genomics Roy J. Carver Biotechnology Center University of Illinois, Urbana-Champaign 330 ERML 1201 W. Gregory Dr. Urbana, IL 61801 USA ph: 217-244-7355 fax: 217-265-5066 e-mail: drnevich at uiuc.edu
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Dear Jenny, Thank you for your reply. I have another question if you would help me. If I sort the MA by MA$genes$ID, the spacing =1, right? spacing has nothing to do with Block, Row, or Column from the gpr file, right? Thanks so much, Tiandao On Tue, 9 Oct 2007, Jenny Drnevich wrote: At 10:38 AM 10/9/2007, Tiandao Li wrote: > Dear Jenny, > > I checked the MA$genes, the following is part of the list > > 1 45Rev2-A03.g > .... > 25 45Rev2-A03.g > ... > 49 45Rev2-A03.g > > 23 rows between 2 replicate spots, so spacing=24? Yes. > Thansk, > > Tiandao > > On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > At 09:19 AM 10/9/2007, Tiandao Li wrote: > >Hello Ido, > > > > Block Row Column ID Name > >519 4 2 17 37A-C02.g P00765 > >531 4 4 17 37A-C02.g P00765 > >513 4 1 17 37A-C02.g P00765 > > > >For 37A-C02.g, I only selected the first couple of genes from the topTable > >list, all 4 replicate spots are printed consecutively in the same column. > > This is not the same as consecutively spacing in ROWS, which is what > the default value of spacing=1 means. Your spacing is probably the > number of spots per row on the array. Check MA$genes to see how many > rows of the data matrix separate your duplicates. > > Jenny > > > > >Thanks for your reply. > > > >Tiandao > > > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > > > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > > Hello, > > > > > > I had arrays with 4 replicate spots per gene. I used limma package for > > > data analysis. > > > > > > > targets > > > > > > SlideNumber FileName Cy3 Cy5 Name > > > Field1 1 13617731 WBM WC Field1 > > > Field2 2 13617730 WBM WC Field2 > > > Field3 3 13617724 WC WBM Field3 > > > Field4 4 13617627 WC WBM Field4 > > > Field5 5 13617626 WBM WC Field5 > > > > > > After read in data, normalization, I used the following codes for > > > within-array replicate spots. > > > > > > design <- modelMatrix(targets, ref="WC") > > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow computation! > > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > method="ls") > > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > method="robust") > > > >Dear Tiandao, > > > >unless the replicate spots are consecutive you have to give a spacing > >argument to indicate what the replicate spots actually are. > >If they are randomly spotted on the array, you would have to rearrange > >them somehow and then use the appropriate spacing. > > > >519 4 2 17 37A-C02.g P00765 > >531 4 4 17 37A-C02.g P00765 > > > >doesn't look as if the replicate spots are one after the other. > > > >HTH best wishes, > >ido > > > >_______________________________________________ > >Bioconductor mailing list > >Bioconductor at stat.math.ethz.ch > >https://stat.ethz.ch/mailman/listinfo/bioconductor > >Search the archives: > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > >_______________________________________________ > >Bioconductor mailing list > >Bioconductor at stat.math.ethz.ch > >https://stat.ethz.ch/mailman/listinfo/bioconductor > >Search the archives: > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > _______________________________________________ > Bioconductor mailing list > Bioconductor at stat.math.ethz.ch > https://stat.ethz.ch/mailman/listinfo/bioconductor > Search the archives: > http://news.gmane.org/gmane.science.biology.informatics.conductor Jenny Drnevich, Ph.D. Functional Genomics Bioinformatics Specialist W.M. Keck Center for Comparative and Functional Genomics Roy J. Carver Biotechnology Center University of Illinois, Urbana-Champaign 330 ERML 1201 W. Gregory Dr. Urbana, IL 61801 USA ph: 217-244-7355 fax: 217-265-5066 e-mail: drnevich at uiuc.edu
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>Thank you for your reply. I have another question if you would help me. If >I sort the MA by MA$genes$ID, the spacing =1, right? spacing has nothing >to do with Block, Row, or Column from the gpr file, right? Correct - the spacing is taken from the order in the MA object and does not rely on the Block, Row & Column info. Be careful about sorting your MA object - as I said before, other spots such as buffers and blanks may appear more than 4 times, and may mess up your ordered sets of 4. You should always double check to make sure the outcome of sorting is what you intended it to be. Jenny >Thanks so much, > >Tiandao > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > >At 10:38 AM 10/9/2007, Tiandao Li wrote: > > Dear Jenny, > > > > I checked the MA$genes, the following is part of the list > > > > 1 45Rev2-A03.g > > .... > > 25 45Rev2-A03.g > > ... > > 49 45Rev2-A03.g > > > > 23 rows between 2 replicate spots, so spacing=24? > >Yes. > > > > Thansk, > > > > Tiandao > > > > On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > > At 09:19 AM 10/9/2007, Tiandao Li wrote: > > >Hello Ido, > > > > > > Block Row Column ID Name > > >519 4 2 17 37A-C02.g P00765 > > >531 4 4 17 37A-C02.g P00765 > > >513 4 1 17 37A-C02.g P00765 > > > > > >For 37A-C02.g, I only selected the first couple of genes from the topTable > > >list, all 4 replicate spots are printed consecutively in the same column. > > > > This is not the same as consecutively spacing in ROWS, which is what > > the default value of spacing=1 means. Your spacing is probably the > > number of spots per row on the array. Check MA$genes to see how many > > rows of the data matrix separate your duplicates. > > > > Jenny > > > > > > > > >Thanks for your reply. > > > > > >Tiandao > > > > > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > > > > > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > > > Hello, > > > > > > > > I had arrays with 4 replicate spots per gene. I used limma package for > > > > data analysis. > > > > > > > > > targets > > > > > > > > SlideNumber FileName Cy3 Cy5 Name > > > > Field1 1 13617731 WBM WC Field1 > > > > Field2 2 13617730 WBM WC Field2 > > > > Field3 3 13617724 WC WBM Field3 > > > > Field4 4 13617627 WC WBM Field4 > > > > Field5 5 13617626 WBM WC Field5 > > > > > > > > After read in data, normalization, I used the following codes for > > > > within-array replicate spots. > > > > > > > > design <- modelMatrix(targets, ref="WC") > > > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow > computation! > > > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > method="ls") > > > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > method="robust") > > > > > >Dear Tiandao, > > > > > >unless the replicate spots are consecutive you have to give a spacing > > >argument to indicate what the replicate spots actually are. > > >If they are randomly spotted on the array, you would have to rearrange > > >them somehow and then use the appropriate spacing. > > > > > >519 4 2 17 37A-C02.g P00765 > > >531 4 4 17 37A-C02.g P00765 > > > > > >doesn't look as if the replicate spots are one after the other. > > > > > >HTH best wishes, > > >ido > > > > > >_______________________________________________ > > >Bioconductor mailing list > > >Bioconductor at stat.math.ethz.ch > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > >Search the archives: > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > >_______________________________________________ > > >Bioconductor mailing list > > >Bioconductor at stat.math.ethz.ch > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > >Search the archives: > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > _______________________________________________ > > Bioconductor mailing list > > Bioconductor at stat.math.ethz.ch > > https://stat.ethz.ch/mailman/listinfo/bioconductor > > Search the archives: > > http://news.gmane.org/gmane.science.biology.informatics.conductor > >Jenny Drnevich, Ph.D. > >Functional Genomics Bioinformatics Specialist >W.M. Keck Center for Comparative and Functional Genomics >Roy J. Carver Biotechnology Center >University of Illinois, Urbana-Champaign > >330 ERML >1201 W. Gregory Dr. >Urbana, IL 61801 >USA > >ph: 217-244-7355 >fax: 217-265-5066 >e-mail: drnevich at uiuc.edu Jenny Drnevich, Ph.D. Functional Genomics Bioinformatics Specialist W.M. Keck Center for Comparative and Functional Genomics Roy J. Carver Biotechnology Center University of Illinois, Urbana-Champaign 330 ERML 1201 W. Gregory Dr. Urbana, IL 61801 USA ph: 217-244-7355 fax: 217-265-5066 e-mail: drnevich at uiuc.edu
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Dear Jenny, Thanks for your help. I messed up rows of MA with rows of MA$genes$Row. I removed the blanks and controls before furthur analysis. I will sort MA by MA$genes$ID to get consecutive spots, and also double check the result. For the difference between methods of ls and robust in lmFit. I looked throught online help. "ls" for least squares or "robust" for robust regression. >From MASS package's help, "robust" is a robust regression using an M estimator, with or without (?) Tukey's biweight initialized by a specific S-estimator. And "robust" here is a iterated re-weighted least squares (IWLS), how did it iterated? Is this the relation between "ls" and "robust" in lmFit? For me, I am wondering the difference of differentially expressed gene lists using methods of "ls" or "robust" in lmFit. Thanks, Tiandao On Tue, 9 Oct 2007, Jenny Drnevich wrote: >Thank you for your reply. I have another question if you would help me. If >I sort the MA by MA$genes$ID, the spacing =1, right? spacing has nothing >to do with Block, Row, or Column from the gpr file, right? Correct - the spacing is taken from the order in the MA object and does not rely on the Block, Row & Column info. Be careful about sorting your MA object - as I said before, other spots such as buffers and blanks may appear more than 4 times, and may mess up your ordered sets of 4. You should always double check to make sure the outcome of sorting is what you intended it to be. Jenny >Thanks so much, > >Tiandao > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > >At 10:38 AM 10/9/2007, Tiandao Li wrote: > > Dear Jenny, > > > > I checked the MA$genes, the following is part of the list > > > > 1 45Rev2-A03.g > > .... > > 25 45Rev2-A03.g > > ... > > 49 45Rev2-A03.g > > > > 23 rows between 2 replicate spots, so spacing=24? > >Yes. > > > > Thansk, > > > > Tiandao > > > > On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > > At 09:19 AM 10/9/2007, Tiandao Li wrote: > > >Hello Ido, > > > > > > Block Row Column ID Name > > >519 4 2 17 37A-C02.g P00765 > > >531 4 4 17 37A-C02.g P00765 > > >513 4 1 17 37A-C02.g P00765 > > > > > >For 37A-C02.g, I only selected the first couple of genes from the topTable > > >list, all 4 replicate spots are printed consecutively in the same column. > > > > This is not the same as consecutively spacing in ROWS, which is what > > the default value of spacing=1 means. Your spacing is probably the > > number of spots per row on the array. Check MA$genes to see how many > > rows of the data matrix separate your duplicates. > > > > Jenny > > > > > > > > >Thanks for your reply. > > > > > >Tiandao > > > > > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > > > > > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > > > Hello, > > > > > > > > I had arrays with 4 replicate spots per gene. I used limma package for > > > > data analysis. > > > > > > > > > targets > > > > > > > > SlideNumber FileName Cy3 Cy5 Name > > > > Field1 1 13617731 WBM WC Field1 > > > > Field2 2 13617730 WBM WC Field2 > > > > Field3 3 13617724 WC WBM Field3 > > > > Field4 4 13617627 WC WBM Field4 > > > > Field5 5 13617626 WBM WC Field5 > > > > > > > > After read in data, normalization, I used the following codes for > > > > within-array replicate spots. > > > > > > > > design <- modelMatrix(targets, ref="WC") > > > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow > computation! > > > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > method="ls") > > > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > method="robust") > > > > > >Dear Tiandao, > > > > > >unless the replicate spots are consecutive you have to give a spacing > > >argument to indicate what the replicate spots actually are. > > >If they are randomly spotted on the array, you would have to rearrange > > >them somehow and then use the appropriate spacing. > > > > > >519 4 2 17 37A-C02.g P00765 > > >531 4 4 17 37A-C02.g P00765 > > > > > >doesn't look as if the replicate spots are one after the other. > > > > > >HTH best wishes, > > >ido > > > > > >_______________________________________________ > > >Bioconductor mailing list > > >Bioconductor at stat.math.ethz.ch > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > >Search the archives: > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > >_______________________________________________ > > >Bioconductor mailing list > > >Bioconductor at stat.math.ethz.ch > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > >Search the archives: > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > _______________________________________________ > > Bioconductor mailing list > > Bioconductor at stat.math.ethz.ch > > https://stat.ethz.ch/mailman/listinfo/bioconductor > > Search the archives: > > http://news.gmane.org/gmane.science.biology.informatics.conductor > >Jenny Drnevich, Ph.D. > >Functional Genomics Bioinformatics Specialist >W.M. Keck Center for Comparative and Functional Genomics >Roy J. Carver Biotechnology Center >University of Illinois, Urbana-Champaign > >330 ERML >1201 W. Gregory Dr. >Urbana, IL 61801 >USA > >ph: 217-244-7355 >fax: 217-265-5066 >e-mail: drnevich at uiuc.edu Jenny Drnevich, Ph.D. Functional Genomics Bioinformatics Specialist W.M. Keck Center for Comparative and Functional Genomics Roy J. Carver Biotechnology Center University of Illinois, Urbana-Champaign 330 ERML 1201 W. Gregory Dr. Urbana, IL 61801 USA ph: 217-244-7355 fax: 217-265-5066 e-mail: drnevich at uiuc.edu _______________________________________________ Bioconductor mailing list Bioconductor at stat.math.ethz.ch https://stat.ethz.ch/mailman/listinfo/bioconductor Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor
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At 02:52 PM 10/9/2007, Tiandao Li wrote: >Dear Jenny, > >Thanks for your help. I messed up rows of MA with rows of MA$genes$Row. > >I removed the blanks and controls before furthur analysis. I will sort MA >by MA$genes$ID to get consecutive spots, and also double check the result. > >For the difference between methods of ls and robust in lmFit. I looked >throught online help. > >"ls" for least squares or "robust" for robust regression. You didn't finish reading through the help file for lmFit - at the end of the Details section there is more information about the linear model computations, and then after that check out the See Also - 06.LinearModels. If this doesn't help you, then you should find a local statistician to help explain it to you. Cheers, Jenny > From MASS package's help, "robust" is a robust regression using an M >estimator, with or without (?) Tukey's biweight initialized by a specific >S-estimator. > >And "robust" here is a iterated re-weighted least squares (IWLS), how did >it iterated? Is this the relation between "ls" and "robust" in lmFit? > >For me, I am wondering the difference of differentially expressed gene >lists using methods of "ls" or "robust" in lmFit. > >Thanks, > >Tiandao > > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > >Thank you for your reply. I have another question if you would help me. If > >I sort the MA by MA$genes$ID, the spacing =1, right? spacing has nothing > >to do with Block, Row, or Column from the gpr file, right? > >Correct - the spacing is taken from the order in the MA object and >does not rely on the Block, Row & Column info. Be careful about >sorting your MA object - as I said before, other spots such as >buffers and blanks may appear more than 4 times, and may mess up your >ordered sets of 4. You should always double check to make sure the >outcome of sorting is what you intended it to be. > >Jenny > > > > > >Thanks so much, > > > >Tiandao > > > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > >At 10:38 AM 10/9/2007, Tiandao Li wrote: > > > Dear Jenny, > > > > > > I checked the MA$genes, the following is part of the list > > > > > > 1 45Rev2-A03.g > > > .... > > > 25 45Rev2-A03.g > > > ... > > > 49 45Rev2-A03.g > > > > > > 23 rows between 2 replicate spots, so spacing=24? > > > >Yes. > > > > > > > Thansk, > > > > > > Tiandao > > > > > > On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > > > > At 09:19 AM 10/9/2007, Tiandao Li wrote: > > > >Hello Ido, > > > > > > > > Block Row Column ID Name > > > >519 4 2 17 37A-C02.g P00765 > > > >531 4 4 17 37A-C02.g P00765 > > > >513 4 1 17 37A-C02.g P00765 > > > > > > > >For 37A-C02.g, I only selected the first couple of genes from > the topTable > > > >list, all 4 replicate spots are printed consecutively in the > same column. > > > > > > This is not the same as consecutively spacing in ROWS, which is what > > > the default value of spacing=1 means. Your spacing is probably the > > > number of spots per row on the array. Check MA$genes to see how many > > > rows of the data matrix separate your duplicates. > > > > > > Jenny > > > > > > > > > > > > >Thanks for your reply. > > > > > > > >Tiandao > > > > > > > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > > > > > > > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > > > > Hello, > > > > > > > > > > I had arrays with 4 replicate spots per gene. I used limma > package for > > > > > data analysis. > > > > > > > > > > > targets > > > > > > > > > > SlideNumber FileName Cy3 Cy5 Name > > > > > Field1 1 13617731 WBM WC Field1 > > > > > Field2 2 13617730 WBM WC Field2 > > > > > Field3 3 13617724 WC WBM Field3 > > > > > Field4 4 13617627 WC WBM Field4 > > > > > Field5 5 13617626 WBM WC Field5 > > > > > > > > > > After read in data, normalization, I used the following codes for > > > > > within-array replicate spots. > > > > > > > > > > design <- modelMatrix(targets, ref="WC") > > > > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow > > computation! > > > > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > > method="ls") > > > > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > > method="robust") > > > > > > > >Dear Tiandao, > > > > > > > >unless the replicate spots are consecutive you have to give a spacing > > > >argument to indicate what the replicate spots actually are. > > > >If they are randomly spotted on the array, you would have to rearrange > > > >them somehow and then use the appropriate spacing. > > > > > > > >519 4 2 17 37A-C02.g P00765 > > > >531 4 4 17 37A-C02.g P00765 > > > > > > > >doesn't look as if the replicate spots are one after the other. > > > > > > > >HTH best wishes, > > > >ido > > > > > > > >_______________________________________________ > > > >Bioconductor mailing list > > > >Bioconductor at stat.math.ethz.ch > > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > > >Search the archives: > > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > > > >_______________________________________________ > > > >Bioconductor mailing list > > > >Bioconductor at stat.math.ethz.ch > > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > > >Search the archives: > > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > > _______________________________________________ > > > Bioconductor mailing list > > > Bioconductor at stat.math.ethz.ch > > > https://stat.ethz.ch/mailman/listinfo/bioconductor > > > Search the archives: > > > http://news.gmane.org/gmane.science.biology.informatics.conductor > > > >Jenny Drnevich, Ph.D. > > > >Functional Genomics Bioinformatics Specialist > >W.M. Keck Center for Comparative and Functional Genomics > >Roy J. Carver Biotechnology Center > >University of Illinois, Urbana-Champaign > > > >330 ERML > >1201 W. Gregory Dr. > >Urbana, IL 61801 > >USA > > > >ph: 217-244-7355 > >fax: 217-265-5066 > >e-mail: drnevich at uiuc.edu > >Jenny Drnevich, Ph.D. > >Functional Genomics Bioinformatics Specialist >W.M. Keck Center for Comparative and Functional Genomics >Roy J. Carver Biotechnology Center >University of Illinois, Urbana-Champaign > >330 ERML >1201 W. Gregory Dr. >Urbana, IL 61801 >USA > >ph: 217-244-7355 >fax: 217-265-5066 >e-mail: drnevich at uiuc.edu > >_______________________________________________ >Bioconductor mailing list >Bioconductor at stat.math.ethz.ch >https://stat.ethz.ch/mailman/listinfo/bioconductor >Search the archives: >http://news.gmane.org/gmane.science.biology.informatics.conductor
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Dear Jenny, After I sorted MA$M and MA$A by MA$genes$ID separately, do I need to sort MA$genes by MA$genes$ID or not? Many Thanks, Tiandao On Tue, 9 Oct 2007, Jenny Drnevich wrote: >Thank you for your reply. I have another question if you would help me. If >I sort the MA by MA$genes$ID, the spacing =1, right? spacing has nothing >to do with Block, Row, or Column from the gpr file, right? Correct - the spacing is taken from the order in the MA object and does not rely on the Block, Row & Column info. Be careful about sorting your MA object - as I said before, other spots such as buffers and blanks may appear more than 4 times, and may mess up your ordered sets of 4. You should always double check to make sure the outcome of sorting is what you intended it to be. Jenny >Thanks so much, > >Tiandao > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > >At 10:38 AM 10/9/2007, Tiandao Li wrote: > > Dear Jenny, > > > > I checked the MA$genes, the following is part of the list > > > > 1 45Rev2-A03.g > > .... > > 25 45Rev2-A03.g > > ... > > 49 45Rev2-A03.g > > > > 23 rows between 2 replicate spots, so spacing=24? > >Yes. > > > > Thansk, > > > > Tiandao > > > > On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > > At 09:19 AM 10/9/2007, Tiandao Li wrote: > > >Hello Ido, > > > > > > Block Row Column ID Name > > >519 4 2 17 37A-C02.g P00765 > > >531 4 4 17 37A-C02.g P00765 > > >513 4 1 17 37A-C02.g P00765 > > > > > >For 37A-C02.g, I only selected the first couple of genes from the topTable > > >list, all 4 replicate spots are printed consecutively in the same column. > > > > This is not the same as consecutively spacing in ROWS, which is what > > the default value of spacing=1 means. Your spacing is probably the > > number of spots per row on the array. Check MA$genes to see how many > > rows of the data matrix separate your duplicates. > > > > Jenny > > > > > > > > >Thanks for your reply. > > > > > >Tiandao > > > > > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > > > > > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > > > Hello, > > > > > > > > I had arrays with 4 replicate spots per gene. I used limma package for > > > > data analysis. > > > > > > > > > targets > > > > > > > > SlideNumber FileName Cy3 Cy5 Name > > > > Field1 1 13617731 WBM WC Field1 > > > > Field2 2 13617730 WBM WC Field2 > > > > Field3 3 13617724 WC WBM Field3 > > > > Field4 4 13617627 WC WBM Field4 > > > > Field5 5 13617626 WBM WC Field5 > > > > > > > > After read in data, normalization, I used the following codes for > > > > within-array replicate spots. > > > > > > > > design <- modelMatrix(targets, ref="WC") > > > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow > computation! > > > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > method="ls") > > > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > method="robust") > > > > > >Dear Tiandao, > > > > > >unless the replicate spots are consecutive you have to give a spacing > > >argument to indicate what the replicate spots actually are. > > >If they are randomly spotted on the array, you would have to rearrange > > >them somehow and then use the appropriate spacing. > > > > > >519 4 2 17 37A-C02.g P00765 > > >531 4 4 17 37A-C02.g P00765 > > > > > >doesn't look as if the replicate spots are one after the other. > > > > > >HTH best wishes, > > >ido > > > > > >_______________________________________________ > > >Bioconductor mailing list > > >Bioconductor at stat.math.ethz.ch > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > >Search the archives: > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > >_______________________________________________ > > >Bioconductor mailing list > > >Bioconductor at stat.math.ethz.ch > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > >Search the archives: > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > _______________________________________________ > > Bioconductor mailing list > > Bioconductor at stat.math.ethz.ch > > https://stat.ethz.ch/mailman/listinfo/bioconductor > > Search the archives: > > http://news.gmane.org/gmane.science.biology.informatics.conductor > >Jenny Drnevich, Ph.D. > >Functional Genomics Bioinformatics Specialist >W.M. Keck Center for Comparative and Functional Genomics >Roy J. Carver Biotechnology Center >University of Illinois, Urbana-Champaign > >330 ERML >1201 W. Gregory Dr. >Urbana, IL 61801 >USA > >ph: 217-244-7355 >fax: 217-265-5066 >e-mail: drnevich at uiuc.edu Jenny Drnevich, Ph.D. Functional Genomics Bioinformatics Specialist W.M. Keck Center for Comparative and Functional Genomics Roy J. Carver Biotechnology Center University of Illinois, Urbana-Champaign 330 ERML 1201 W. Gregory Dr. Urbana, IL 61801 USA ph: 217-244-7355 fax: 217-265-5066 e-mail: drnevich at uiuc.edu _______________________________________________ Bioconductor mailing list Bioconductor at stat.math.ethz.ch https://stat.ethz.ch/mailman/listinfo/bioconductor Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor
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You should NEVER sort the internal components of your MA list separately! To sort all of them at once, just sort on the MA object itself: MA2 <- MA[order(MA$genes$ID) , ] This will keep all the internal components in the same order. You *really* should start playing around with the objects on your own, and reading through all the help files instead of immediately asking questions to the list. This will help you learn R better, and increase the odds that someone will respond to your question when you really get stuck. Good luck, Jenny At 04:22 PM 10/9/2007, Tiandao Li wrote: >Dear Jenny, > >After I sorted MA$M and MA$A by MA$genes$ID separately, do I need to sort >MA$genes by MA$genes$ID or not? > >Many Thanks, > >Tiandao > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > >Thank you for your reply. I have another question if you would help me. If > >I sort the MA by MA$genes$ID, the spacing =1, right? spacing has nothing > >to do with Block, Row, or Column from the gpr file, right? > >Correct - the spacing is taken from the order in the MA object and >does not rely on the Block, Row & Column info. Be careful about >sorting your MA object - as I said before, other spots such as >buffers and blanks may appear more than 4 times, and may mess up your >ordered sets of 4. You should always double check to make sure the >outcome of sorting is what you intended it to be. > >Jenny > > > > > >Thanks so much, > > > >Tiandao > > > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > >At 10:38 AM 10/9/2007, Tiandao Li wrote: > > > Dear Jenny, > > > > > > I checked the MA$genes, the following is part of the list > > > > > > 1 45Rev2-A03.g > > > .... > > > 25 45Rev2-A03.g > > > ... > > > 49 45Rev2-A03.g > > > > > > 23 rows between 2 replicate spots, so spacing=24? > > > >Yes. > > > > > > > Thansk, > > > > > > Tiandao > > > > > > On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > > > > At 09:19 AM 10/9/2007, Tiandao Li wrote: > > > >Hello Ido, > > > > > > > > Block Row Column ID Name > > > >519 4 2 17 37A-C02.g P00765 > > > >531 4 4 17 37A-C02.g P00765 > > > >513 4 1 17 37A-C02.g P00765 > > > > > > > >For 37A-C02.g, I only selected the first couple of genes from > the topTable > > > >list, all 4 replicate spots are printed consecutively in the > same column. > > > > > > This is not the same as consecutively spacing in ROWS, which is what > > > the default value of spacing=1 means. Your spacing is probably the > > > number of spots per row on the array. Check MA$genes to see how many > > > rows of the data matrix separate your duplicates. > > > > > > Jenny > > > > > > > > > > > > >Thanks for your reply. > > > > > > > >Tiandao > > > > > > > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > > > > > > > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > > > > Hello, > > > > > > > > > > I had arrays with 4 replicate spots per gene. I used limma > package for > > > > > data analysis. > > > > > > > > > > > targets > > > > > > > > > > SlideNumber FileName Cy3 Cy5 Name > > > > > Field1 1 13617731 WBM WC Field1 > > > > > Field2 2 13617730 WBM WC Field2 > > > > > Field3 3 13617724 WC WBM Field3 > > > > > Field4 4 13617627 WC WBM Field4 > > > > > Field5 5 13617626 WBM WC Field5 > > > > > > > > > > After read in data, normalization, I used the following codes for > > > > > within-array replicate spots. > > > > > > > > > > design <- modelMatrix(targets, ref="WC") > > > > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow > > computation! > > > > > fit <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > > method="ls") > > > > > fit2 <- lmFit(MA, design, ndups=4, correlation=corfit$consensus, > > > > > method="robust") > > > > > > > >Dear Tiandao, > > > > > > > >unless the replicate spots are consecutive you have to give a spacing > > > >argument to indicate what the replicate spots actually are. > > > >If they are randomly spotted on the array, you would have to rearrange > > > >them somehow and then use the appropriate spacing. > > > > > > > >519 4 2 17 37A-C02.g P00765 > > > >531 4 4 17 37A-C02.g P00765 > > > > > > > >doesn't look as if the replicate spots are one after the other. > > > > > > > >HTH best wishes, > > > >ido > > > > > > > >_______________________________________________ > > > >Bioconductor mailing list > > > >Bioconductor at stat.math.ethz.ch > > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > > >Search the archives: > > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > > > >_______________________________________________ > > > >Bioconductor mailing list > > > >Bioconductor at stat.math.ethz.ch > > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > > >Search the archives: > > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > > _______________________________________________ > > > Bioconductor mailing list > > > Bioconductor at stat.math.ethz.ch > > > https://stat.ethz.ch/mailman/listinfo/bioconductor > > > Search the archives: > > > http://news.gmane.org/gmane.science.biology.informatics.conductor > > > >Jenny Drnevich, Ph.D. > > > >Functional Genomics Bioinformatics Specialist > >W.M. Keck Center for Comparative and Functional Genomics > >Roy J. Carver Biotechnology Center > >University of Illinois, Urbana-Champaign > > > >330 ERML > >1201 W. Gregory Dr. > >Urbana, IL 61801 > >USA > > > >ph: 217-244-7355 > >fax: 217-265-5066 > >e-mail: drnevich at uiuc.edu > >Jenny Drnevich, Ph.D. > >Functional Genomics Bioinformatics Specialist >W.M. Keck Center for Comparative and Functional Genomics >Roy J. Carver Biotechnology Center >University of Illinois, Urbana-Champaign > >330 ERML >1201 W. Gregory Dr. >Urbana, IL 61801 >USA > >ph: 217-244-7355 >fax: 217-265-5066 >e-mail: drnevich at uiuc.edu > >_______________________________________________ >Bioconductor mailing list >Bioconductor at stat.math.ethz.ch >https://stat.ethz.ch/mailman/listinfo/bioconductor >Search the archives: >http://news.gmane.org/gmane.science.biology.informatics.conductor Jenny Drnevich, Ph.D. Functional Genomics Bioinformatics Specialist W.M. Keck Center for Comparative and Functional Genomics Roy J. Carver Biotechnology Center University of Illinois, Urbana-Champaign 330 ERML 1201 W. Gregory Dr. Urbana, IL 61801 USA ph: 217-244-7355 fax: 217-265-5066 e-mail: drnevich at uiuc.edu
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Further to Jenny's suggestion, > summary(MA$genes$ID) should tell you if the blanks/controls etc are there in more than 4 copies. hth, a > -----Original Message----- > From: bioconductor-bounces at stat.math.ethz.ch > [mailto:bioconductor-bounces at stat.math.ethz.ch] On Behalf Of > Jenny Drnevich > Sent: 09 October 2007 18:57 > To: Tiandao Li > Cc: bioconductor at stat.math.ethz.ch > Subject: Re: [BioC] HELP! lmFit and duplicateCorrelation > > > > >Thank you for your reply. I have another question if you > would help me. > >If I sort the MA by MA$genes$ID, the spacing =1, right? spacing has > >nothing to do with Block, Row, or Column from the gpr file, right? > > Correct - the spacing is taken from the order in the MA object and > does not rely on the Block, Row & Column info. Be careful about > sorting your MA object - as I said before, other spots such as > buffers and blanks may appear more than 4 times, and may mess up your > ordered sets of 4. You should always double check to make sure the > outcome of sorting is what you intended it to be. > > Jenny > > > > > >Thanks so much, > > > >Tiandao > > > >On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > >At 10:38 AM 10/9/2007, Tiandao Li wrote: > > > Dear Jenny, > > > > > > I checked the MA$genes, the following is part of the list > > > > > > 1 45Rev2-A03.g > > > .... > > > 25 45Rev2-A03.g > > > ... > > > 49 45Rev2-A03.g > > > > > > 23 rows between 2 replicate spots, so spacing=24? > > > >Yes. > > > > > > > Thansk, > > > > > > Tiandao > > > > > > On Tue, 9 Oct 2007, Jenny Drnevich wrote: > > > > > > At 09:19 AM 10/9/2007, Tiandao Li wrote: > > > >Hello Ido, > > > > > > > > Block Row Column ID Name > > > >519 4 2 17 37A-C02.g P00765 > > > >531 4 4 17 37A-C02.g P00765 > > > >513 4 1 17 37A-C02.g P00765 > > > > > > > >For 37A-C02.g, I only selected the first couple of genes > from the > > > >topTable list, all 4 replicate spots are printed > consecutively in > > > >the same column. > > > > > > This is not the same as consecutively spacing in ROWS, > which is what > > > the default value of spacing=1 means. Your spacing is > probably the > > > number of spots per row on the array. Check MA$genes to > see how many > > > rows of the data matrix separate your duplicates. > > > > > > Jenny > > > > > > > > > > > > >Thanks for your reply. > > > > > > > >Tiandao > > > > > > > >On Tue, 9 Oct 2007, Ido M. Tamir wrote: > > > > > > > >On Tuesday 09 October 2007 01:20, Tiandao Li wrote: > > > > > Hello, > > > > > > > > > > I had arrays with 4 replicate spots per gene. I used limma > > > > > package for data analysis. > > > > > > > > > > > targets > > > > > > > > > > SlideNumber FileName Cy3 Cy5 Name > > > > > Field1 1 13617731 WBM WC Field1 > > > > > Field2 2 13617730 WBM WC Field2 > > > > > Field3 3 13617724 WC WBM Field3 > > > > > Field4 4 13617627 WC WBM Field4 > > > > > Field5 5 13617626 WBM WC Field5 > > > > > > > > > > After read in data, normalization, I used the following codes > > > > > for within-array replicate spots. > > > > > > > > > > design <- modelMatrix(targets, ref="WC") > > > > > corfit <- duplicateCorrelation(MA, design, ndups=4) # A slow > > computation! > > > > > fit <- lmFit(MA, design, ndups=4, > correlation=corfit$consensus, > > > > > method="ls") > > > > > fit2 <- lmFit(MA, design, ndups=4, > correlation=corfit$consensus, > > > > > method="robust") > > > > > > > >Dear Tiandao, > > > > > > > >unless the replicate spots are consecutive you have to give a > > > >spacing argument to indicate what the replicate spots > actually are. > > > >If they are randomly spotted on the array, you would have to > > > >rearrange them somehow and then use the appropriate spacing. > > > > > > > >519 4 2 17 37A-C02.g P00765 > > > >531 4 4 17 37A-C02.g P00765 > > > > > > > >doesn't look as if the replicate spots are one after the other. > > > > > > > >HTH best wishes, > > > >ido > > > > > > > >_______________________________________________ > > > >Bioconductor mailing list > > > >Bioconductor at stat.math.ethz.ch > > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > > >Search the archives: > > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > > > >_______________________________________________ > > > >Bioconductor mailing list > > > >Bioconductor at stat.math.ethz.ch > > > >https://stat.ethz.ch/mailman/listinfo/bioconductor > > > >Search the archives: > > > >http://news.gmane.org/gmane.science.biology.informatics.conductor > > > > > > _______________________________________________ > > > Bioconductor mailing list > > > Bioconductor at stat.math.ethz.ch > > > https://stat.ethz.ch/mailman/listinfo/bioconductor > > > Search the archives: > > > http://news.gmane.org/gmane.science.biology.informatics.conductor > > > >Jenny Drnevich, Ph.D. > > > >Functional Genomics Bioinformatics Specialist > >W.M. Keck Center for Comparative and Functional Genomics > >Roy J. Carver Biotechnology Center > >University of Illinois, Urbana-Champaign > > > >330 ERML > >1201 W. Gregory Dr. > >Urbana, IL 61801 > >USA > > > >ph: 217-244-7355 > >fax: 217-265-5066 > >e-mail: drnevich at uiuc.edu > > Jenny Drnevich, Ph.D. > > Functional Genomics Bioinformatics Specialist > W.M. Keck Center for Comparative and Functional Genomics > Roy J. Carver Biotechnology Center > University of Illinois, Urbana-Champaign > > 330 ERML > 1201 W. Gregory Dr. > Urbana, IL 61801 > USA > > ph: 217-244-7355 > fax: 217-265-5066 > e-mail: drnevich at uiuc.edu > > _______________________________________________ > Bioconductor mailing list > Bioconductor at stat.math.ethz.ch > https://stat.ethz.ch/mailman/listinfo/biocondu> ctor > Search the > archives: > http://news.gmane.org/gmane.science.biology.informatics.conductor > -- The Wellcome Trust Sanger Institute is operated by Genome Research Limited, a charity registered in England with number 1021457 and a company registered in England with number 2742969, whose registered office is 215 Euston Road, London, NW1 2BE.
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