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Tiling Arrays
Madelaine GogolProgrammer [email protected]
Norman PavelkaPostdocRong Li [email protected]
Technology and Methods SeminarMarch 29, 2007
http://wiki/research/MadelaineGogol
Tiling Arrays - Overview
• What is a tiling array?• What can I do with it?
– ChIP-chip– CGH– Expression
• Which tiling arrays are available for my experiments?– in-house yeast tiling array (design details)– Agilent– Affymetrix
• How will we analyze and visualize the data?
What is a Tiling Array?
Gene 1
Probe 1
A microarray with many probes distributed in an evenly spaced way across an entire genome.
What can I do with tiling arrays?• Map the transcriptome
– what’s being expressed?
• ChIP-chip– where are proteins binding?
• CGH– what are the differences in genome structure?
• Other possibilities– Map the methylome– Genome resequencing– Polymorphism discovery
In-house yeast tiling array (YOGie)
• Covers the yeast genome
• Just printed
• resolution ~ 250 bases
• freely available
Operon Probe Set
• 6307 probes, length 70• Designed one per ORF, near 3’ end.• YOG arrays (yeast oligonucleotide)• YBOX: 3072 new probes
Intergenic Probe Set: Design
• Design target: yeast Intergenic regions• original goal: leave no area > 500 uncovered
Gene 1 Gene 2
Array Oligo
Selector (AOS)
Fasta format 140-mer sequences tiling the intergenic regions
9,405 70-mer sequences from the intergenic regions
Intergenic Probe Set
• 9405 70-mer probes• No region greater than 360 left uncovered• ~ 220 bases between probes on average• Chromosome 3 completely tiled• YOGi arrays (YOG+intergenic)
5’ probe set: Design
• Goal: – fill in gaps left by operon set in 5’ region of gene– leave no region > 500 without a probe
• Targets: ORF regions 5’ of operon probe
Gene 1
operon probe
3’5’
Array Oligo
Selector (AOS)
5’ probe set: Reduction
• Too many probes– reduce to 6666 or less (budget and printing constraints)
• Probes within 260 bases of eachother – winnowed
• Tm
• binding energy
• number of matches to genome
Gene 1
operon probe
3’5’
5’ probe set
• 6,512 70-mer probes• tiles the region of each gene between the operon
probe and the 5' end. • YOGie arrays (YOG+intergenic+enhanced)
In-house yeast tiling array: YOGie
Together, the operon, intergenic, and 5’ sets make up our homemade yeast tiling array
• Freely available• Also includes tight tiles of
– all centromeres– 7 sub-genomic regions 10-20 kb
Agilent Tiling Arrays
ChIP-on-chip Arabidopsis Whole Genome C. elegans Whole GenomeDrosophila Whole GenomeHuman CpG Island Human ENCODE 244KHuman Promoter Mouse Promoter Yeast Whole Genome 4 x 44K Yeast Whole Genome 244K Zebrafish Expanded Promoter Zebrafish Proximal Promoter Custom ChIP-on-chip
Oligo aCGH Human Genome 244KHuman Genome 105KHuman Genome 44KMouse Genome 244KMouse Genome 105KMouse Genome 44K
244k
105k
44k
15k
1
2
4
8
$400
$640
$720
$800
$320
$180
$100
per slide
Agilent Tiling Arrays: formats and cost
per hyb
$400
Agilent Custom Array Design
• Take an agilent microarray design• Remove some probes• Put in your own probes
– Design using agilent’s web application, earray.
• You can also design everything from scratch
http://www.affymetrix.com/support/technical/byproduct.affx?cat=exparrays
Affymetrix Tiling Arrays
Arabidopsis Tiling 1.0R Array C.elegans Tiling 1.0R Array Chromosome 21/22 1.0 Array Set Chromosome 21/22 2.0R Array Drosophila Tiling 1.0R Array ENCODE01 1.0 ArrayHuman Genome Arrays +Mouse Genome Arrays +S. cerevisiae Tiling 1.0R ArrayS. pombe Tiling 1.0FR Array
Cost ~ 500$ per array, so 500$ per hyb.
Summary of Tiling Arrays (only yeast shown)
Type #spots per array
probe size
resolution per slide cost
per array cost
customizable?
Homemade YOGie arrays
26,000 70 250 between
0 0 some effort
Agilent 44K 44,000 (4) 60 160 between
$720 $180 easily
Agilent 244K 244,000 60 overlap by 18 $400 $400 easily
Affy 3,200,000(pm/mm)
25 overlap by 20 $500 $500 no
Tiling array data analysis: still an adventure
• Affy– TAS (Tiling Analysis Software)
• Agilent – ChIP Analytics & CGH Analytics
• Other– genome browsers, R packages, other people’s software,
Do-it-yourself, perl, statistical models
Data Visualization: Genome Browsers: UCSC
http://wiki/research/Bioinformatics/GenomeBrowsers
http://genome.ucsc.edu/cgi-bin/hgTracks?db=sacCer1&hgt.customText=http://research.stowers-institute.org/mcm/YOGie1.bed
Data Analysis: Average Gene analysis
Profile of binding across an average gene
0 20 40 60 80
0.0
0.2
0.4
0.6
AcH4/WT
positions in average gene and surrounding intergenic regions
ave
rag
ed
exp
ress
ion
ra
tios
http://wiki/research/Microarray/AverageGeneAnalysis
Summary
• Tiling arrays– CHip-chip, CGH, expression
• Which ones are available– In-house, Agilent, Affy
• Data analysis
• Future...
Other Future Genomic Technology
454 and Solexa/Illumina “Next Generation” sequencing• “Sequence everything in the tube”• Shares some things with tiling arrays
– even more unbiased– vast quantities of data– analysis methods are being developed
5th floorBuilding 2
N
Bioinformatics
Microarray
Allison Karin Brian ChrisMe
Thanks!
Microarray
Bioinformatics
All the labs that use microarrays!
Bing LiWorkman lab
Jennifer BuppJasperson lab
Norman PavelkaRong Li Lab
Technology & Methods Technology & Methods SeminarSeminar
““Adventures in Electron Adventures in Electron Microscopy”Microscopy”
Rhonda AllenRhonda AllenHistologyHistology
Thursday, April 26th, 1:00 p.m.Thursday, April 26th, 1:00 p.m.ClassroomClassroom
(1(1stst floor, Administration Building) floor, Administration Building)
Schedule with abstracts and previous presentation slides can be found on:Schedule with abstracts and previous presentation slides can be found on:K:\Weekly Seminar Schedule\Thursday -- Technology & MethodsK:\Weekly Seminar Schedule\Thursday -- Technology & Methods
Information regarding previous seminars can be found at:Information regarding previous seminars can be found at:http://research.stowers-institute.org/wiw/external/Seminars/index.htmhttp://research.stowers-institute.org/wiw/external/Seminars/index.htm