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What Is Microarray

What Is Microarray. A new powerful technology for biological exploration Parallel High-throughput Large-scale Genomic scale. Microarray. Replication. Transcription. Translation. DNA. RNA. Protein. Reverse transcription. DNA. ATATCGGCATCAGTCGATCGATCATCGATCGAT. mRNA.

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What Is Microarray

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  1. What Is Microarray • A new powerful technology for biological exploration • Parallel • High-throughput • Large-scale • Genomic scale

  2. Microarray

  3. Replication Transcription Translation DNA RNA Protein Reverse transcription DNA ATATCGGCATCAGTCGATCGATCATCGATCGAT mRNA UAUAGCCGUAGUCAGCUAGCUAGUAGCUAGCUA cDNA ATATCGGCATCAGTCGATCGATCATCGATCGAT

  4. What Is Microarray • Different Approaches

  5. Affymetrix • Probe Array (Photolithography) • Synthesis of probe • Hybridization

  6. Stanford Approach • Use robot to spot glass slides • Able to measure qualitatively relative expression levels of genes • Differential expression by use of simultaneous, two-color fluorescence hybridisation • Cheaper with DIY ($60,000) • Also called home-made system

  7. Life Cycle(cont.) Cy3 (green) Cy5 (red)

  8. Arrayer Hardware Scanner Software Procedures • Preparation • Target DNA (reference and test samples) • Slides • Reaction(Droplet or Pin Spotting) • Hybridization • Scanning • Analysis • Image processing • Data mining • Modeling

  9. Instruments • Arrayer • Scanner • Laser Confocal Microscope

  10. Arrayer

  11. Scanner • GenPix 4000

  12. Problems – by direct view Anti-probe Locally high background Spot overlap Precipitate Locally low signal Comet-tails (donut hole)

  13. Problems –related wit IP • Local problems • Spot position variation • Spot shape and size irregularity • Spot Overlapping • Global problems • High background • Low signal • White-noise/Speckle effect (Contamination)

  14. Array Alignment Good Alignment Bad Alignment

  15. Spot Finding • Grid System Determination • Manual • Semiautomatic • Automatic • Spot Segmentation • Space-based • Intensity-based • Frequency-based • Hybrid approach

  16. Grid System Determination

  17. Spot Segmentation

  18. Spot Segmentation(cont.)

  19. Applications • Gene expression studies • Gene function for cell state change in various conditions • Disease Diagnosis • Pathogen Analysis

  20. Applications(Cont.) • Drug Discovery • identify appropriate molecular targets for therapeutic intervention • monitor changes in gene expression in response to drug treatments • Trageted Drug Treatment

  21. Example Images – E. Coli

  22. Example Images – C. Elegan

  23. ….. Gene index 1.75 Log(R/G) …..

  24. Log(R/G) Log(R/G) Log(R/G) ….. ….. ….. ….. ….. ….. …

  25. M experiments N gene …..

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