Bioinformatics, 2012.11.15
Gene Expression Profiling by Microarray
Chun-Ju Chang, [email protected] of Food ScienceCollege of Life SciencesNational Taiwan Ocean University
2Griffin & Shockcor . Nature Reviews Cancer 2004, 4:551.
High Throughput Gene Discovery• Solution for genomics study
3Sutliff J. Science 2001, 291:1224.
Gene chip (DNA chip, DNA microarray) Microarray technology evolved
from Southern blotting, where fragmented DNA is attached to a substrate and then probed with a known gene or fragment.
Nucleic Acids Res. 1992, 20:1679.
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Schematic of microarray analysisSchematic of microarray analysis
AnalysisAnalysisAnalysisAnalysis
Quality Quality measurementmeasurement Pre-ProcessingPre-Processing
FaileFailedd
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Outline• Microarray platforms• Experimental design
– Sources of variability – Sample size and replication
• Data acquisition and preprocessing– Normalization– Quality control
• Data analysis– Partitional clustering– Functional annotation– Pathway analysis
• Gene expression databases
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Microarray platforms 1
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Microarray platforms 2
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Affymetrix
Miller & Tang. Microbiol Rev. 2009,22:611.
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Illumina
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250,000 probes/bead
Experimental design 1Sources of variation in a microarray experiment :• Manufacturing of arrays• Generation of biological sample
– Genetic and environmental factors– Pooled or individual samples – Randomization
• Technical variation – Preprocessing : RNA extraction, labeling, etc.– Protocolization of the processing steps
• Processing of samples – Obtaining image– “Biological replicates”, “technical replicates”
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Experimental design 2Sample size and replication:
• 4 types of experimental designs– Completely randomized treatment-control design: each
measurement is considered independent– Matched-pairs design– Multiple treatment design having an independent
treatment effect– Randomized block design
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Data acquisition and preprocessingCommon normalization strategies• Total intensity normalization• Normalization using regression techniques• Normalization using ratio statistics
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Data acquisition and preprocessingQuality control• From the Microarray Gene Expression Data (MGED) Society; presently
named Functional Genomics Data (FGED) Society• MIAME (Minimum Information About a Microarray Experiment)
standards for data reporting– Spotted cDNA and oligonucleotide arrays– Experimental design: number of replicates, samples used– Preparation and labeling– Hybridization procedures and parameters– Measurement data and specifications
• Microarray Gene Expression Markup Language (MAGE-ML)• ArrayExpress microarray database
– Universal data-presentation platform
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Functional Genomics Data (FGED) Society
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MicroArray Quality Control (MAQC) project
16Ji H & Davis RW. Nat Biotechnol 2006, 24:1112-3.
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Scatter plotHierarchical Trees
Pi Chart
K-MeansK-Means
Venn diagramVenn diagram
Partitional clustering by K-Means
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cluster centers, prototypes
反覆疊代
Functional annotation by GO (Gene Ontology)
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EMBL-EBI
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Hands-on Practice
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