Post on 20-Mar-2018
Agilent SurePrint G3 CGH+SNP
Microarray Platform
Your Complete Solution For Cytogenetic Research
For Research Use Only. Not for use in diagnostic procedures.
Complete workflow
• SurePrint G3 CGH+SNP microarrays
• Copy number changes and LOH/UPD on a single array
• Highest quality copy-number data
• Processing reagents and Automation
• Instruments and reagents optimized for Agilent microarrays, for greater
consistency of results
• Increased throughput with Bravo Automated Liquid Handling Platform
• Agilent CytoGenomics 1.5 analysis software
• Designed specifically for cytogeneticists
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For Research Use Only. Not for use in diagnostic procedures.
SUREPRINT G3 CGH+SNP
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For Research Use Only. Not for use in diagnostic procedures.
Breakthrough dual CGH+SNP microarray
• Single microarray that offers high resolution aberration calls in
addition to LOH/UPD (loss of heterozygosity/uniparental
disomy) detection
– No need to choose between high resolution copy number data and
LOH/UPD or alternatively run two separate microarrays
• Addition of SNP probes to the Agilent CGH SurePrint G3 CGH
arrays enables the detection of blocks of LOH/UPD
– CGH calls not affected by addition
of SNP content, high quality
copy-number data remains
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For Research Use Only. Not for use in diagnostic procedures.
UPD: uniparental disomy
• Both members of a chromosome pair or segments of a chromosome pair are
inherited from one parent
• UPD can result in an abnormal phenotype when the chromosomes involved are
imprinted, such that only the maternal or paternal allele of the pair is active
Remark: parental
samples needed
to detect
heterodisomy
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For Research Use Only. Not for use in diagnostic procedures.
LOH: loss of heterozygosity
• UPD in tumor cells is often referred to as acquired UPD or copy neutral LOH
(cnLOH)
• Common in both hematologic and solid tumors
Blood, Vol. 108, Issue 5, 1733-1743, September 1, 2006
normal cell
trisomic cell
LOH of entire
chromosome
LOH of arm
LOH of
part of
armmonosomic cell
interstitial regions of LOH
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For Research Use Only. Not for use in diagnostic procedures.
Measuring SNPs using restriction enzymes
• A subset of probes on a CGH array
measures SNPs in parallel to copy
number, on a single array
• Genotype SNPs using restriction
digestion (Alu/Rsa)
• Measure the copy number of one
allele at each SNP site relative to a
known reference
• Regions of LOH are located by
finding genomic regions with a
statistically significant scarcity of
heterozygous calls
• ~5-10 Mb LOH/UPD resolution
across the entire genome
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For Research Use Only. Not for use in diagnostic procedures.
DATA
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For Research Use Only. Not for use in diagnostic procedures.
Call rate at 95% confidence: >95%
Twelve samples hybridized against a single HapMap reference sample
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For Research Use Only. Not for use in diagnostic procedures.
Call accuracy: >99%
Three HapMap samples hybridized against a single HapMap
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For Research Use Only. Not for use in diagnostic procedures.
Detection of UPDe.g. NA20409: complete paternal isodisomy on chr. 15
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SNP
data
CGH
data
No heterozygous SNPs
Copy-neutral LOH or UPD
Only TWO states of alleles appear:
1) 0 uncut copies
2) 2 uncut copies
Both copies were
inherited from the same
parent – UPD
BB
AB
AA
For Research Use Only. Not for use in diagnostic procedures.
Confirmation of Copy Number Changese.g. NA09209: hemizygous deletion on chr. 17
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SNP
data
CGH
dataDeletion at the
beginning of p-arm
Hemizygous deletion
TWO states of alleles appear (only one copy allele):
1) cut copy of the allele (0 copy)
2) uncut copy of the allele (1 copy)
B
A
For Research Use Only. Not for use in diagnostic procedures.
Confirmation of Copy Number Changes e.g. NA04592: trisomy of chr. 21
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Amplification
FOUR or more states
Each state corresponds to 0, 1, 2, 3 (or more) copies of the uncut allele
Amplification of chr. 21
SNP
data
CGH
data
BBB
ABB
AAB
AAA
For Research Use Only. Not for use in diagnostic procedures.
Detection of Consanguinitye.g. NA06231: child from consanguineous parents
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Extended segments of
homozygosity found
throughout the
genome
SNP
data
CGH
data
BB
AB
AA
For Research Use Only. Not for use in diagnostic procedures.
Excellent correlation between Agilent and ILMN
93 LOH regions discovered in 3 consanguineous samples run on Agilent
CGH+SNP and Illumina 610 quad platforms
1
10
100
1 10 100Agile
nt C
GH
+S
NP
data
: L
OH
siz
e in M
b
Illumina Beadchip data: LOH size in Mb
LOH region size on Illumina and Agilent platforms
10.7 and 5.2 Mb regions
detected on Agilent but not
Illumina
5.3, 3.7, and 3.6 Mb
regions detected on
Illumina but not Agilent
r = 0.9952
Data generated at Baylor College of Medicine
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For Research Use Only. Not for use in diagnostic procedures.
AGILENT CYTOGENOMICS 1.5
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For Research Use Only. Not for use in diagnostic procedures.
Streamline data analysis for Cytogenetic labs
• Simple workflow to analyze samples processed on Agilent SurePrint G3
CGH and CGH+SNP microarrays.
• Fast 5-minute data analysis for a normal sample.
• Convenient input and output support connection to LIMS.
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Sample info
Laboratory LIMSCyto report
Triage
& sign off
Run data
analysis
For Research Use Only. Not for use in diagnostic procedures.
Powerful algorithms for calling CNC, LOH and UPD
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Feature
Extraction
ADM-1
ADM-2
SNP CN & LOH
Log2 ratio of
intensities to call CNCs
No. of uncut alleles
to call LOH/UPD
For Research Use Only. Not for use in diagnostic procedures.
Sample triage
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Not approved for use in diagnostic
procedures
Preloaded tracks:
DGV-CNV, OMIM
Custom tracks
Auto updated
tracks based on
aberration
classification in
CytoGenomics
Database
Add notes to
aberrations
View aberrations
in UCSC
genome browser
Classify
aberrations
Showing number
of samples with
similar aberrations
in database
Suppress
aberrations
Right click on Classification allows query for
overlapping aberrations in database: results will
be shown as new track
Link out from gene
to external
sources: DGV,
OMIM, Entrez
For Research Use Only. Not for use in diagnostic procedures.
Additional features in Agilent CytoGenomics 1.5
• Interval classification and database queries for similar aberrations
• Workflow Automation Mode for automatic processing of TIFF images and
report generation
• Upload data to Cartagenia bench
• Supports multiple versions of genome build and ability to lock into a specific
genome build
• Links to OMIM and OMIM morbid entries from aberrant intervals
• Multiple levels of user privilege settings
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Download CytoGenomics free trial @ https://earray.chem.agilent.com/earray/
For Research Use Only. Not for use in diagnostic procedures.
PUBLICATIONS
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For Research Use Only. Not for use in diagnostic procedures.
Evaluation of high-resolution array platforms
Agilent arrays:
• show the highest dynamic
range
• slope of the measurements
closest to the theoretical
values
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Plot of measured log2 ratios compared to theoretical
values
Kresse et al. BMC Res Notes. 2010;3:223
For Research Use Only. Not for use in diagnostic procedures.
• More probe density does not mean more array resolution
• Even with half the number of probes the Agilent 1M array has:
– better exon/gene coverage
– lower DLRSD
– less signal variation
– higher copy-number detection rate
• Higher DLR values in SNP6.0
reduces the resolution of
abnormality detection
– SNP6.0 unable to call a focal gain
on PAX5 identified by 8 and 31
probes using Agilent 244K and
Agilent 1M arrays, respectively
Agilent 244K, 1M, Affymetrix SNP6.0 comparison
Braggio et al. AACR 101st Annual Meeting, April 2010
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For Research Use Only. Not for use in diagnostic procedures.
DNA copy number array technologies assessed
• Agilent was the best performing platform and for the SNP-CGH platforms,
Affymetrix tended to outperform Illumina.
• Nimblegen failed to detect numerous aberrations that were clear in the other
platforms even when probes were tiled in the region of interest.
• Nimblegen exhibits 2-4 fold greater variance amongst replicate probes and
variances an order of magnitude greater for replicate array comparisons.
Clarity of signal Agilent > Affymetrix > Illumina > Nimblegen
Curtis et al. BMC Genomics. 2009;10:588
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For Research Use Only. Not for use in diagnostic procedures.
Automation of the CGH sample preparation with
the Bravo Automated Liquid Handling Platform
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• Agilent Automation Solutions (formerly Velocity 11) Bravo platform was
successfully used in the WTCCC 19,000 sample landmark CNV study
• 19,000 samples in 19 weeks
For Research Use Only. Not for use in diagnostic procedures.
PRODUCT INFORMATION
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For Research Use Only. Not for use in diagnostic procedures.
SurePrint G3 CGH+SNP Products
TypePart
NumberProduct Description
CGH
probe
focus
# slides
per kit
SurePrint G3
Catalog
G4842A SurePrint G3 CGH+SNP Array Kit 2x400K Genes and
exons5
G4890A SurePrint G3 CGH+SNP Array Kit 4x180K ISCA 3
SurePrint G3
Custom
G4882A SurePrint G3 Custom CGH+SNP 1x1M Custom 1
G4883A SurePrint G3 Custom CGH+SNP 2x400K Custom 1
G4884A SurePrint G3 Custom CGH+SNP 4x180K Custom 1
G4885A SurePrint G3 Custom CGH+SNP 8x60K Custom 1
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For Research Use Only. Not for use in diagnostic procedures.
Agilent Catalog ISCA CGH+SNP Microarray
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CGH probesSNP probes
60K SNP probes ~5-10 mb LOH
4x180K SurePrint G3 CGH+SNP kit P/N G4890A
For Research Use Only. Not for use in diagnostic procedures.
Design your Custom ISCA CGH+SNP Microarray
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Visit: https://www.agilent.com/genomics/earray
• Use the ISCA targeted
regions probe groups
• Add your own content in
eArray
For Research Use Only. Not for use in diagnostic procedures.
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For Research Use Only. Not for use in diagnostic procedures.