Biochemistry 412

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Biochemistry 412 2005 5 April Lecture DNA
Microarrays
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Microarrays DNA Arrays Protein Arrays
Other
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Microarrays DNA Arrays Protein Arrays
Other
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Microarrays Have Led to an Explosion in mRNA
Profiling Studies
Stolovitky (2003) Curr. Opin. Struct. Biol. 13,
370.
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Two Main Types of DNA Microarrays
Grünenfelder Winzeler (2002) Nature Rev. Genet.
3, 653.
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Lockhart Winzeler (2000) Nature 405, 827.
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Affymetrix Gene Chips - In Situ Synthesis
Note 4N masks required to make an array of
oligonucleotides, each of length N.
Pease et al (1994) Proc. Natl. Acad. Sci. U.S.A.
91, 5022.
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Note this is the photolabile blocking group,
X, indicated schematically in Figure 1.
Pease et al (1994) Proc. Natl. Acad. Sci. U.S.A.
91, 5022.
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Key feature known oligo sequence at each
address on the chip.
Lipshutz et al (1999) Nature Genet. (suppl.) 21,
20.
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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 Disadva
ntage - big initial investment less
standardization
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Note Not all arrays have to be on chips! -
Illumina, Inc.
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Caveat.Caveat.Caveat!! Results from the
different DNA microarray methods dont always
agree! And results for mRNA
abundance differences dont always agree with
protein abundance data!
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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
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Applications of DNA Microarrays
Genotyping mRNA profiling and
transcriptome analysis Genome analysis
(cancer and evolutionary studies)
Genome-wide splicing analyses Etc.
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Applications of DNA Microarrays
Genotyping mRNA profiling and
transcriptome analysis Genome analysis
(cancer and evolutionary studies)
Genome-wide splicing analyses Etc.
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Comparative mRNA Analyses Using Microarrays
Bryant et al (2004) Lancet Infect. Disease 4, 100.
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Technical Proficiency Experimental Design are
Key to Reproducibility
Lockhart Barlow (2001) Nature Rev. Neurosci. 2,
63.
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However, if your technique isnt good enough, the
data must be normalized...
Forster et al (2003) J. Endocrinol. 178, 195.
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Microarray data Analysis of a hypothetical time
course of mRNA abundances for several genes
Leung Cavalieri (2003) Trends in Genetics 19,
649.
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Note caloric restriction gene chip experiment
w/ rats.
Ref Lee et al (1999) Science 285, 1390.
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Lee et al (1999) Science 285, 1390.
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Common Genes Induced in Immature Dendritic Cells
in Response to Infection with Diverse Pathogens
Huang et al (2001) Science 294, 870.
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Bassett et al (1999) Nature Genet. (suppl.) 21,
51.
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Applications of DNA Microarrays
Genotyping mRNA profiling and
transcriptome analysis Genome analysis
(cancer and evolutionary studies)
Genome-wide splicing analyses Etc.
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Microarrys Can Also be Used to Analyze
Chromosomal Rearrangments
CGH - Comparative genomce hybridization LOH -
Loss of heterozygosity
Albertson Pinkel (2003) Human Molec. Genet. 12,
R145.
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Applications of DNA Microarrays
Genotyping mRNA profiling and
transcriptome analysis Genome analysis
(cancer and evolutionary studies)
Genome-wide splicing analyses Etc.
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Microarrays can be used to detect tissue-specific
alternative splicing
Blue - tissue 1 Purple - tissue 2 Yellow - pooled
sample (average)
Le et al (2004) Nucleic Acids Research 32, e180.
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Microarray probe elements specifically designed
to detect alternative splicing
Barrass Beggs (2003) Trends in Genetics 19, 295.
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Applications of DNA Microarrays
Genotyping mRNA profiling and
transcriptome analysis Genome analysis
(cancer and evolutionary studies)
Genome-wide splicing analyses Etc.
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RNA as a repository of past generations genetic
information??
Weigel Jürgens (2005) Nature 434, 443.
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