1. Development, Implementation and Testing of a DNA Microarray Test Suite Ehsanul Haque Mentors: Dr. Cecilie Boysen Dr. Jim Breaux ViaLogy Corp.

2. Outline Purpose Purpose Background Background Datasets Datasets Algorithms Algorithms Results Results Conclusion Conclusion

3. Purpose To design tests to assess the results from Different microarray data analysis services Different microarray data analysis services VMAxS, GCOS VMAxS, GCOS Different updates of the same data analysis service Different updates of the same data analysis service VMAxS_V1, VMAxS_V2, VMAxS_V3, VMAxS_V4 VMAxS_V1, VMAxS_V2, VMAxS_V3, VMAxS_V4 Different background correction, normalization and summarization methods Different background correction, normalization and summarization methods VMAxS, PLIER, RMA, dCHIP VMAxS, PLIER, RMA, dCHIP Different technologies of microarray data analysis Different technologies of microarray data analysis Affymetrix, ABI, Agilent, GE Healthcare, Illumina Affymetrix, ABI, Agilent, GE Healthcare, Illumina

4. Background VMAxS RA Report (Resonance Amplitude) Probe level signal data 11-20 probe per gene 1 expression value per gene (if from Affymetrix system) VMAxS Signal Detection via QRI DAT or raw image file Only raw data files accepted Quantum Resonance Interferometry (QRI) – Active Signal Processing

5. From image to gene level data Raw pixel level data VMAxS/GCOS Probe level data VMAxS/MAS5.0/PLIER/RMA Background correction Normalization Summarization (PM/MM) Gene level data Image analysis

6. Datasets.DAT File Method D Method C Method B Method A Each Dataset stems from the same raw AffymetrixGeneChip.dat files GeneChip family Human 2A: 54676 genes 3 Replicates * 8 Dilutions 24 Samples

7. Test Suite [R scripting] Master Function Function CV (Coefficient of Variance) User File Function RA (Relative Accuracy) Function DEG (Differential Expressed Gene) Other Output File list & parameters

8. Algorithm Function RA (Rela. Accu.) Function TABLE.IN Using a loop, calls function TABLE.IN to work on the User File list of input datasets Calls other functions, parses different formats of data- sets and provides function RA with the required table. Method AMethod B Method CMethod D 0.497897086864 0.394078146688 0.33660070237 0.211275056257 Slope 0.497897086864 0.394078146688 0.33660070237 0.211275056257 User File File list & parameters Calculates Mean & SD Calculates Slope and Ave. of mean intensities Plots Slope vs. Average of mean intensity Plots Conc. vs. Mean Intensity Creates a list of Median Slope Replaces zero values

9. CV Plot Three Replicates A, B & C

10. CV Plot With fitted line

11. 8 Dilutions Mean Intensity vs. Concentration 8 Dilutions Mean Intensity vs. Concentration

12. Slope vs. Mean Intensity

13. Median Slope

14. Results Tests Dataset processed by Method A Method B Method C Method D CV0.1170554806340.1719023236030.07271146285460.100852274894 Rel. Accu. 0.4978970868640.3940781466880.336600702370.211275056257 DEG More

15. Conclusion Developed and implemented two tests in R Developed and implemented two tests in R CV & RA (relative accuracy) CV & RA (relative accuracy) Tested 2 X 4 datasets Tested 2 X 4 datasets Different parameters of the same analysis method Different parameters of the same analysis method Different normalization methods Different normalization methods More tests are needed to draw a better conclusion about the data. More tests are needed to draw a better conclusion about the data.

16. Reference & Acknowledgment References: Zhijin Wu el al, A Model Based Background Adjustment of Oligonucleotide Expression Arrays, Johns Hopkins University, Dept of Biostatistics Working Papers, 2004, paper 1 Zhijin Wu el al, A Model Based Background Adjustment of Oligonucleotide Expression Arrays, Johns Hopkins University, Dept of Biostatistics Working Papers, 2004, paper 1 Irizarry et al, Exploration, Normalization, and Summaries of High Density Oligonucleotide Array Probe Level Data, Biostatistics (2003), 4, 2, pp. 249-264 Irizarry et al, Exploration, Normalization, and Summaries of High Density Oligonucleotide Array Probe Level Data, Biostatistics (2003), 4, 2, pp. 249-264 Cope et al, A Benchmark of Affymetrix GeneChip Expression Measures, Bioinformatics, Vol. 1 no. 1 2003, pp. 1-10 Cope et al, A Benchmark of Affymetrix GeneChip Expression Measures, Bioinformatics, Vol. 1 no. 1 2003, pp. 1-10Acknowledgements: Dr. Cecilie Boysen Dr. Cecilie Boysen Dr. Jim Breaux Dr. Jim Breaux - Vialogy - Vialogy Southern California Bioinformatics Summer Institute (SoCalBSI) Southern California Bioinformatics Summer Institute (SoCalBSI) National Institute of Health (NIH) National Institute of Health (NIH)