Biochemistry 4122005
5 April Lecture
DNA Microarrays
Microarrays
• DNA Arrays
• Protein Arrays
• Other
Microarrays
• DNA Arrays
• Protein Arrays
• Other
Stolovitky (2003) Curr. Opin. Struct. Biol. 13, 370.
Microarrays Have Led to an Explosion in mRNA Profiling Studies
Grünenfelder & Winzeler (2002) Nature Rev. Genet. 3, 653.
Two Main Types of DNA Microarrays
Lockhart & Winzeler (2000) Nature 405, 827.
Pease et al (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 5022.
Note: 4N masks required tomake an array of oligonucleotides,each of length N.
Affymetrix Gene Chips - In Situ Synthesis
Pease et al (1994) Proc. Natl. Acad. Sci. U.S.A. 91, 5022.
Note: this is the photolabile blockinggroup, “X”, indicated schematicallyin Figure 1.
Lipshutz et al (1999) Nature Genet. (suppl.) 21, 20.
Key feature: known oligosequence at each “address”on the chip.
In situ synthesized (Affymetrix) microarrays:
Advantage - massive coverage of sequences possible
Disadvantage - expensive and not easily customized
Spotted arrays (oligonucleotides or dsDNA):
Advantage - flexibility and cost; can be “home-made”
Disadvantage - big initial investment; less standardization
Note:Not all arrays have to be on chips…!
- Illumina, Inc.
Caveat….Caveat….Caveat…!!
• Results from the differentDNA microarray methodsdon’t always agree!
• And results for mRNA abundancedifferences don’t always agreewith protein abundance data!
Stages in the Life Cycle of a Technological Innovation
1. Unveiling (first publication)
2. Hype (lots of follow-on publications, meetings, venture capital, etc.)
3. Disillusionment (more paper opportunities!)
4. Promise eventually fulfilled (years later)
5. Enters routine use
Applications of DNA Microarrays
• Genotyping• mRNA profiling and
“transcriptome” analysis• Genome analysis (cancer and
evolutionary studies)• Genome-wide splicing analyses• Etc.
Applications of DNA Microarrays
• Genotyping• mRNA profiling and
“transcriptome” analysis• Genome analysis (cancer and
evolutionary studies)• Genome-wide splicing analyses• Etc.
Comparative mRNA Analyses Using Microarrays
Bryant et al (2004)Lancet Infect. Disease4, 100.
Technical Proficiency & Experimental Design are Key to Reproducibility
Lockhart & Barlow (2001)Nature Rev. Neurosci. 2, 63.
However, if your technique isn’t good enough, the data must be “normalized”...
Forster et al (2003)J. Endocrinol. 178, 195.
Microarray data: Analysis of a hypothetical time course of mRNA
abundances for several genes
Leung & Cavalieri (2003) Trends in Genetics 19, 649.
Ref: Lee et al (1999) Science 285, 1390.
Note: caloric restriction gene chip experiment w/ rats.
Lee et al (1999) Science 285, 1390.
Huang et al (2001) Science 294, 870.
Common Genes Induced in Immature Dendritic Cells in Response to Infection with Diverse Pathogens
Bassett et al (1999) Nature Genet. (suppl.) 21, 51.
Applications of DNA Microarrays
• Genotyping• mRNA profiling and
“transcriptome” analysis• Genome analysis (cancer and
evolutionary studies)• Genome-wide splicing analyses• Etc.
Microarrys Can Also be Used to Analyze Chromosomal Rearrangments
Albertson & Pinkel (2003) Human Molec. Genet. 12, R145.
CGH - “Comparative genomce hybridizationLOH - “Loss of heterozygosity”
Applications of DNA Microarrays
• Genotyping• mRNA profiling and
“transcriptome” analysis• Genome analysis (cancer and
evolutionary studies)• Genome-wide splicing analyses• Etc.
Microarrays can be used to detect tissue-specific alternative splicing
Blue - tissue 1Purple - tissue 2Yellow - pooled sample (average) Le et al (2004) Nucleic Acids Research 32, e180.
Microarray probe elements specifically designed to detect alternative splicing
Barrass & Beggs (2003)Trends in Genetics 19, 295.
Applications of DNA Microarrays
• Genotyping• mRNA profiling and
“transcriptome” analysis• Genome analysis (cancer and
evolutionary studies)• Genome-wide splicing analyses• Etc.
RNA as a repository of past generations’ genetic information??
Weigel & Jürgens (2005) Nature 434, 443.