Statistics for Human Genetics and Molecular BiologyLecture 22: Introduction to Microarray Technology
Dr. Yen-Yi Ho ([email protected])
Oct 28, 2015
1/34
Objectives of Lecture 22
I Types of ArraysI Expression Microarrays
I General Experiment ProtocolI Hybridization, Target, Probe, Probe SetI Commercial MicroarraysI Common Issues
I aCGH
I ChIP-chip
2/34
Microarrays
I Microarrays (or just arrays) allow the simultaneousmeasurement of the mRNA of thousands of genes (geneexpression)
I Why “micro”? Measuring things on a small scale means lessmRNA is necessary to be extracted
I The simultaneous measurement is important, so that variousfactors/conditions are all common to the array, and relativegene expression can be measured with less variability
I Still microarray? Microarray is still widely used because oflower costs, easier experimental procedure and moreestablished analysis methods.
I Similar problems are presented in newer technologies such asRNA-seq, and similar statistical approaches can be applied.
3/34
Types of Arrays
1. DNA
• SNP arrays:genotype studies
• array CGH:DNA Copy Number
• ChIP-chip:TF binding sites
2. mRNA
• DNA microarrays:custom array,Affymetrix, Agilent,Illumina
• SAGE
3. Proteins
• 2-D electrophoresis• Maldi-Tof mass spec
4/34
Types of Arrays
1. DNA
• SNP arrays:genotype studies
• array CGH:DNA Copy Number
• ChIP-chip:TF binding sites
2. mRNA
• custom array,Affymetrix, Agilent,Illumina
• SAGE
3. Proteins
• 2-D electrophoresis• Maldi-Tof mass spec
5/34
Co$t
MicorarrayAffymetrix Cost: $425/per arrayIllumina BeadChip Cost: $ 80 ∼ $250 /per strip × 12 = $960 ∼$3,000 /chip
Next-Generation SequencingIllumina ∼$2,000 per samples (for 1-7 samples)
Source: University of Minnesota Genomics Center
6/34
Steps in Obtaining Gene Expression Data Using Microarray
7/34
8/34
9/34
Target
10/34
Probes
11/34
Microarray Experiment Procedure
12/34
Platforms that dominate market
I Affymetrix
I Agilent
I Illumina
I NimbleGen
13/34
Affymetrix GeneChip R©
source: Affymatrix
14/34
Affymetrix GeneChip R©
source: Affymatrix15/34
Affymetrix GeneChip R©
16/34
Probe Set
A probe set is a collection of probes designed to interrogate agiven sequence.
17/34
PM versus MM
18/34
Affymetrix GeneChip R© Experiment Protocol
19/34
Illumina BeadArrays
• Each silica bead is 3 µ in diameter
• 700,000 copies of same probe sequence attached to each bead
• May have more than one bead for a particular gene
20/34
Illumina BeadArrays
• RefSeq BeadChip (left) 8 arrays per chip, 1 strip= 1 array
• Whole Genome (right) 6 arrays per chip, 2 strip= 1 array
21/34
Commons Issues
I Background: print-tip, plate, print-order, spatial effects
I Between arrays: batches, plates, cross platform comparison,experiment protocols
I Within arrays: background noise, intensity dependent effect
22/34
Spatial Effect
23/34
Spatial Effect
24/34
25/34
26/34
Types of Arrays
1. DNA
• SNP arrays:genotype studies
• array CGH:DNA Copy Number
• ChIP-chip:TF binding sites
2. mRNA
• DNA microarrays:cDNA, Affymetrix,Agilent
• SAGE
3. Proteins
• 2-D electrophoresis• Maldi-Tof mass spec
27/34
Array-comparative genomic hybridization (aCGH)
Nature Education1(1). Microarray-based Comparative Genomic Hybridization (aCGH)
28/34
array CGH
Brian and Development 31(6):629:633.
29/34
Types of Arrays
1. DNA
• SNP arrays:genotype studies
• array CGH:DNA Copy Number
• ChIP-chip:TF binding sites
2. mRNA
• DNA microarrays:cDNA, Affymetrix,Agilent
• SAGE
3. Proteins
• 2-D electrophoresis• Maldi-Tof mass spec
30/34
ChIP-chipChromatin immunoprecipitation (”ChIP”) with microarraytechnology (”chip”)
31/34
ChIP-chip
32/34
Types of Arrays
1. DNA
• SNP arrays:genotype studies
• array CGH:DNA Copy Number
• ChIP-chip:TF binding sites
2. mRNA
• DNA microarrays:cDNA, Affymetrix,Agilent
• SAGE
3. Proteins
• 2-D electrophoresis• Maldi-Tof mass spec
33/34
Readings
Bioconductor Case Studies: Chapter 1, 2, 3 (page 1 -45).
34/34