Hi Lilian,

The ontology analysis can be difficult and even with human genes you can get
significant problems.
DAVID used to take the GeneIDs as input as well, the problem with that is that
many transcripts on the array can be unclassified.
However, if you can convert the Agilent IDs to GeneIDs that can improve your
current results. Since you are working with a well characterized biological
model, monocytes and macrophages, it should also improve your chances of getting
sensible ontology analysis.

Resourcer at Harward, http://compbio.dfci.harvard.edu/tgi/cgi-bin/magic/r1.pl is
a great way to get your annotations, but they do not yet offer Agilent Bovine
annotation there. you might want to still have a look and contact them to see if
they can help. Another ontology analysis resource that you may want to have a
look at is Sorin Draghici's Onto-Tools http://vortex.cs.wayne.edu/projects.htm.

I hope that helps a bit if you have any questions please contact me,

Agnieszka

-----------------
Agnieszka Lichanska, PhD
Staff Scientist
TessArae, LLC
http://www.tessarae.com





________________________________
From: Lilian <lijoli@...>
To: microarray@yahoogroups.com
Sent: Tuesday, 30 June, 2009 6:26:33 PM
Subject: [microarray-group] Microarray analysis advise needed!!! Thanks





Hello all!

I have ran an microarray experiment analysis comparing monocytes from blood to
macrophages from endometrium from pregnant cows. I used the bovine agilent chip
4x44K, and my experimental design is showed below. I tried to used the
R-Bioconductor to analyze my data but there is no annotation package for the
bovine agilent array, also I tried the LimmaGUI, but I could not use the
LimmaGui because of my technical replications were read as a biological
replications. So, I used the JMP genomics to analyze my data. My raw data was
previous normalized within array using the LOWESS normalization and in the JMP I
performed the quantile normalization to normalize the data between arrays. I
used the proc ANOVA to do the stats and find the differential regulated genes.
PROC ANOVA
Model: Animal, Replicate, Tissue
Fixed Effects: Tissue, Replicate, Replicate*Tissue
Random effects: Animal, Animal*Tissue, Animal*Replicate, Animal*Tissue*
Replicate

I used the FDR correction fixed at 0.01 the genes were considered differentially
expressed (DE) had p-value<0.05 and at least 2-fold increased ou decreased
expression.

I have got around DE 1,200 genes and now I am working on the ontology analysis.
I have a problem because the agilent ID is not read in most of the softwares for
gene ontology and in general I get only 300 genes annotated out of over 600
genes that I analyze using the DAVID. Do you have any comments on that? Is that
normal not get the ontology analysis for more than a half genes?

I am really new on microarray analysis and I am working by myself, it has been a
big challenge for me to do it, so I really appreciate any comments that you
might have.

Thanks in advance

Lilian Oliveira

Lilian Oliveira DVM Ms
Ph.D. Candidate
Animal Molecular and Cell Biology
University of Florida
Phone/Fax: (352) 392-5590/392- 5595

Replicate Tissue Channel
1 1 Blood Cy3
1 1 Endo CY5
2 1 Blood Cy3
2 1 Endo CY5
3 1 Blood Cy3
3 1 Endo CY5
4 1 Blood Cy3
4 1 Endo CY5
1 2 Endo Cy3
1 2 Blood Cy5
2 2 Endo Cy3
2 2 Blood Cy5
3 2 Blood Cy3
3 2 Endo Cy5
4 2 Endo Cy3
4 2 Blood Cy5

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