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Introduction to microarray technology

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Outline

A little background - molecular level

Types of microarrays

cDNA arrays

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What a microarray does

Detect and measure gene expression

at the mRNA level

Detection of protein level?

Other uses?

Many different types of microarrays

(called platforms)

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Common to all platforms

All have a many biomolecules fixed onto a well-defined surface in known positions (“probes”)

low density 100s (e.g. protein antibodies), medium density 1000s to 10s of 1000s (e.g. cDNA arrays), high-density is 100s to 1000s of 1000s, i.e.millions (e.g. short oligonucleotide arrays).

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Common to all platforms

All have a many biomolecules fixed onto a well-defined surface in known positions (“probes”)

low density 100s (e.g. protein antibodies), medium density 1000s to 10s of 1000s (e.g. cDNA arrays), high-density is 100s to 1000s of 1000s, i.e.millions (e.g. short oligonucleotide arrays).

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Basic Aspects

a) coupling biomolecules (“probes”) to a platform; b) preparing samples for detection; c) hybridization; d) scanning; and e) analyzing the data. Obviously we’re interested in e), but without good knowledge of a) to d), that would be dangerous.

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cDNA arrays: history

cDNA microarrays have evolved from Southern blots, with clone libraries gridded out on nylon membrane filters being an important and still widely used intermediate. Things took off with the introduction of non-porous solid supports, such as glass - these permitted miniaturization - and fluorescence based detection.

Currently, up to about 30,000 cDNAs are spotted onto a microscope slide.

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The cDNA and short (25 bp) oligo technologies in brief. Long (60-75 bp) oligo arrays are more like the cDNA ones

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Nucleic acid hybridization: here DNA-RNA

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Rudiments, completed

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Cartoon version: Before labelling

Array 1 Array 2

Sample 1 Sample 2

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Before Hybridization

Array 1 Array 2

Sample 1 Sample 2

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After Hybridization

Array 1 Array 2

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Quantification

Array 1 Array 2

4 2 0 3 0 4 0 3

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The cDNA and short (25 bp) oligo technologies in brief. Long (60-75 bp) oligo arrays are more like the cDNA ones

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Discuss

1. You want to measure the expression levels of 20,000 genes. How would you arrange them on a microarray (in which order)?

2. The description says you are measuing this in two samples at the same time. You are taking relative measurements. What is a reason for doing this?