Question: [maSigPro] How to analyze 2-colour microarrays?
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gravatar for Maciej Jończyk
9.5 years ago by
Maciej Jończyk720 wrote:
Dear List Members, First - my experiment I have 7 time points and two treatments (cold and control). I used two-colour microarrays, each RNA was hybridized with a common reference (time zero). I've searched list's archive and read the tutorial but still I can't figure out how can I analyze this experiment (two-colour with common reference) in maSigPro. I know that I have to split my data (long time series). I'd like to make two variants of analysis: 1 - Normalization in limma, analysis in maSigPro. 2 - ASCA-genes, then analysis in maSigPro (normalization?). My questions are: (1) - Should I use log-ratio (time_point to time_zero on each microarray) in analysis? (2) - How experimental design file should look like in this case? I've read about ASCA-genes methodology, which can be used as a preprocessing method to subsequent analysis in maSigPro. I've found codes but I don't know how the results from ASCA-genes can be passed to maSigPro. Should I normalize the data before using ASCA-genes? Help will be very appreciated. Best Regards, Maciej Jo?czyk, MSc Department of Plant Molecular Ecophysiology Institute of Plant Experimental Biology Faculty of Biology, University of Warsaw 02-096 Warszawa, Miecznikowa 1 ___________________________________ NOCC, http://nocc.sourceforge.net -- This email was Anti Virus checked by Astaro Security Gateway. http://www.astaro.com
normalization limma masigpro • 611 views
ADD COMMENTlink modified 9.5 years ago by Mª José Nueda90 • written 9.5 years ago by Maciej Jończyk720
Answer: [maSigPro] How to analyze 2-colour microarrays?
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gravatar for Mª José Nueda
9.5 years ago by
Spain
Mª José Nueda90 wrote:
Dear Maciej, You can apply maSigPro with 7 time-points without splitting the data. The method is going to work well. You must split the data if the obtained models don't fit very well. Splitting the data is not implemented automatically in the maSigPro R-program and you have to elaborate a more complicate design. We have developed a web-service where we have implemented maSigPro and other tools http://sea.bioinfo.cipf.es/. We have tried to simplify things. If you want apply maSigPro easily you can use it. We are checking that all work well. If you use it you could send your opinion. You can visit the "help" to see examples: data and results. Regarding your question, it is not necesary to use log-ratio (time_point to time_zero on each microarray) to apply maSigPro. On the other hand ASCA-genes is also a methodology to detect significant genes. We are developing an strategy based on ASCA to eliminate the bacth effect. This is a preprocessing technique but it is not a normalization method. We haven't published anything about this yet. Where did you hear about it?When we finish all the things we will add in SEA web-service also this tool. Best wishes, Mar?a. ----- Original Message ----- From: "Maciej Jo?czyk" <mjonczyk@biol.uw.edu.pl> To: <bioconductor at="" stat.math.ethz.ch=""> Cc: "Maciej Jo?czyk" <mjonczyk at="" biol.uw.edu.pl=""> Sent: Tuesday, April 20, 2010 1:48 PM Subject: [BioC] [maSigPro] How to analyze 2-colour microarrays? > Dear List Members, > > First - my experiment > > I have 7 time points and two treatments (cold and control). > I used two-colour microarrays, each RNA was hybridized with a common > reference (time zero). > > I've searched list's archive and read the tutorial but still I can't > figure out how can I analyze this experiment (two-colour with common > reference) in maSigPro. > I know that I have to split my data (long time series). > > I'd like to make two variants of analysis: > > 1 - Normalization in limma, analysis in maSigPro. > > 2 - ASCA-genes, then analysis in maSigPro (normalization?). > > My questions are: > > (1) - Should I use log-ratio (time_point to time_zero on each > microarray) in analysis? > (2) - How experimental design file should look like in this case? > > I've read about ASCA-genes methodology, which can be used as a > preprocessing method to subsequent analysis in maSigPro. I've found > codes but I don't know how the results from ASCA-genes can be passed to > maSigPro. > > Should I normalize the data before using ASCA-genes? > > Help will be very appreciated. > Best Regards, > > Maciej Jo?czyk, MSc > Department of Plant Molecular Ecophysiology > Institute of Plant Experimental Biology > Faculty of Biology, University of Warsaw > 02-096 Warszawa, Miecznikowa 1 > > > > ___________________________________ > NOCC, http://nocc.sourceforge.net > > > > > -- > This email was Anti Virus checked by Astaro Security Gateway. > http://www.astaro.com > > _______________________________________________ > 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
ADD COMMENTlink written 9.5 years ago by Mª José Nueda90
Dear Mar?a, thank you for response Maria Jose Nueda <mj.nueda at="" ua.es=""> nadawca : > Dear Maciej, > > You can apply maSigPro with 7 time-points without splitting the data. > The > method is going to work well. You must split the data if the obtained > models > don't fit very well. Splitting the data is not implemented > automatically in > the maSigPro R-program and you have to elaborate a more complicate > design. Ok, I'll try analyze my data without splitting and I'll see how it works. > We have developed a web-service where we have implemented maSigPro and > other > tools http://sea.bioinfo.cipf.es/. We have tried to simplify things. > If you > want apply maSigPro easily you can use it. We are checking that all > work > well. If you use it you could send your opinion. You can visit the > "help" to > see examples: data and results. I want to try analysis in R first, but I'll look on this site. > Regarding your question, it is not necesary to use log-ratio > (time_point to > time_zero on each microarray) to apply maSigPro. This question was inspired by the form of design file. In the "maSigPro-tutorial.pdf" you gave examples. I've thought that there is only one row per array (in other case name of each array was written two times - for sample labeled with cy3 and cy5, respectively), so I thought that data must be in log-ratio form. What is the correct form of design file for data in log2(fluorescence) form? > On the other hand ASCA-genes is also a methodology to detect > significant > genes. We are developing an strategy based on ASCA to eliminate the > bacth > effect. This is a preprocessing technique but it is not a > normalization > method. We haven't published anything about this yet. Where did you > hear > about it? I've read about it in your doctoral thesis, which I 've found on the internet. I've found it an interesting method, and the tesis is very inspiring. Now I want to use ASCA-genes as a preprocessing technique for subsequent analysis in maSigPro, and I wonder how could I transport objects between this two packages. Best wishes, Maciej ___________________________________ NOCC, http://nocc.sourceforge.net
ADD REPLYlink written 9.5 years ago by Maciej Jończyk720
I forgot to mention that in my experiment I've used reference design with dye swap (two labelling of reference with cy3 and samples with cy5 and two reverse). Maybe it is important in view of my questions. Maria Jose Nueda <mj.nueda at="" ua.es=""> nadawca : > You can apply maSigPro with 7 time-points without splitting the data. > The > method is going to work well. You must split the data if the obtained > models > don't fit very well. Splitting the data is not implemented > automatically in > the maSigPro R-program and you have to elaborate a more complicate > design. So I should use regression model of degree 6, or am I wrong? Thank you again, Best Regards, Maciej Jo?czyk, MSc Department of Plant Molecular Ecophysiology Institute of Plant Experimental Biology Faculty of Biology, University of Warsaw 02-096 Warszawa, Miecznikowa 1 ___________________________________ NOCC, http://nocc.sourceforge.net
ADD REPLYlink written 9.5 years ago by Maciej Jończyk720
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