DNA Microarray

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DNA Microarray
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Microarray Printing
96-well-plate (PCR Products)
384-well print-plate
Microarray
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Differential Expression

• Each cell contains a complete copy of the
  organisms genome
• Cells are of many different types and state
• e.g. blood, nerve, skin cells, etc
• What makes the cells different ?
• Differential gene expression, i.e., when, where
  and in what quantity each gene is expressed
• On average, 40 of our genes are expressed at
  any given time

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Functional genomics

• The various genome projects have yielded the
  complete DNA sequences of many organisms.
• e.g. human, mouse, yeast, fruitfly, etc.
• Human 3 billion base-pairs, 30-40 thousand
  genes.
• Challenge go from sequence to function,
• i.e., define the role of each gene and
  understand how the genome functions as a whole.

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Central Dogma

• The expression of the genetic information stored
  in the DNA Molecule occurs in two stages
• --transcription, during which DNA is
  transcribed into mRNA
• --translation, during which mRNA is
  translated to produce a protein.
• DNA mRNA Protein

cDNA Arrays
Tissue Arrays
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The Central Dogma of Molecular Biology

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Microarray Hybridization
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Microarray Gene Expression Image
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A Better Look
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Image Analysis Data Visualization
Cy5 Cy3
log2
Cy3
Cy5
Experiments
8 4 2 fold 2 4 8
Underexpressed Overexpressed
Genes
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SpotList
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Ovarian Tumor Study M. Schaner
Samples that should Cluster together do not
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Data Normalization
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Pool of Cell Lines
Tumor
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Such biases have consequences

• Plotting the frequency of un-normalized
  intensities reveals the differential effect
  between the two c hannels.

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How do we deal with this?

• Normalization
• In general, an assumption is made that the
  average gene does not change.
• You must understand your experiment and data to
  judge whether that assumption is a good one.
• Usually true for gene expression experiments, but
  not necessarily for aCGH or chromatin IP.
• Generally true for large arrays, but not for
  small " boutique" arrays.

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Normalization The R-I Plot

• Data may have an intensity-dependent structure.
• Plot log2(R/G) vs. log10(RG) to reveal this
• Reveals
• variance in log ratios is greater at lower
  intensities.
• distribution may not be centered around zero.

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Normalization Loess
log2(R/G)
log10(RG)
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Cluster Analysis

• Cell Cycle example( Spellman 1988)

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Overview of the Cell Cycle

• Purpose
• To create two new cells by dividing one original
  cell

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Cell Cycle Key Concepts

• All parts of original cell must be replicated and
  split between new cells
• Each step must occur in precise manner and timing
  for successful cycle, and is strictly regulated
• mRNA and proteins for cell cycle genes are found
  at varying levels at different points of the
  cycle
• Mutations causing malfunction in regulation can
  result in cancer

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Yeast Cell Cycle
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Cell Cycle Basic Description
http//www.bmb.psu.edu/courses/biotc489/notes/cycl
e.jpg
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Cells grow out of synchrony.
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