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# Differential Expression Testing #
#                                 #
#     ***   README  ***           #
#                                 #
#   Ricardo Verdugo               #
#   The Jackson Laboratory        #
#   October 3rd, 2009             #
#   Bar Harbor, Maine             #
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Title: Differential expression in a C57BL/6J-chrYA congenic strain versus its C57BL/6J background genotype in
myocardial cells.


Chromosome Y (chrY) from the mouse strain C57BL/6J (ChrYC57) was substituted for that of A/J mice (ChrYA).
Cardiomyocytes from the resulting "chromosome substitution" C57BL/6J-chrYA strain were smaller than that of
their C57BL/6J counterparts. In reverse, when chrYA from A/J mice was substituted for that of chrYC57,
cardiomyocytes from the resulting A/J-chrYC57 strain are larger than in their A/J counterparts. Genome-wide
gene expression in cardiomyocytes was compared between these strains using the Illumina Mouse-ref8 BeadChip.
The effects of strain and post-pubertal testosterone levels were tested by ANOVA of a factorial design with
four experimental groups by genotype and castration treatment: ChrYA/intact, ChrYA/castrated, C57/intact,
C57/castrated. Four animals per group were tested.

References
For more information about the experimental samples:
 Llamas, Bastien, Ricardo Verdugo, Gary Churchill, and Christian Deschepper. 2009.
 Chromosome Y variants from different inbred mouse strains are linked to differences in the
 morphologic and molecular responses of cardiac cells to postpubertal testosterone. BMC
 Genomics 10, no. 1 (April 7): 150. doi:10.1186/1471-2164-10-150.

For informations about the analysis of this data:
 Verdugo, Ricardo A., Christian F. Deschepper, Gloria Munoz, Daniel Pomp, and Gary A. Churchill.
 2009. Importance of randomization in microarray experimental designs with Illumina platforms.
 Nucl. Acids Res. 37, no. 17 (September): 5610-8. doi:10.1093/nar/gkp573.

GEO database, record GSE15354.
http://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE15354


Data analysis
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An R script is included to produce a full differential expression analysis.

All necessary data is included. However, in order to successfully run the script, you need to have R
installed, which can be obtained from http://www.r-project.org, as well as the following R packages:

 * maanova
 * affy
 * preprocessCore
 * limma
 
You can find them along with installation instructions at http://www.bioconductor.org
 
Once they are installed, you should be able to run the analysis by reading the
 'YChrom-DE.R' R script from within R.
 
 There are several ways to accomplish this:
 
 * In Windows:
    1) Click "File/Change dir..."
    2) Browse to the directory containing this README and the YChrom-DE.R script
    3) Click "Source R code..." and Accept
    
 * In any system, once you are in the R command line window, type:
    > setwd('root_path') # Replace root_path for the path to the location of this README
    > source('YChrom-DE.R')

This will run a full data analysis workflow:
    1) Readin raw data
    2) Quality control
    3) Data Normalization
    4) Probe filtering
    4) Testing differential expression
    5) Exporting annotated results
    6) Produce some venn diagram plots
    
    
Prepared by
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Ricardo A. Verdugo


Contact Person
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Gary Churchill  at gary.churchill@jax.org