Download Quantitative Analysis of Methylation with Single

APPLICATION BRIEF
Targeted Methylation
Analysis
Quantitative Analysis of Methylation with Single-Base Resolution
Discover epigenetic variations associated with complex human diseases
Demonstrated Performance: DNA
Methylation in Cancer Samples — Kiel
University Study
In a study led by a team of researchers
at Kiel University, the RainDance platform was used to analyze DNA methylation status in patients with colon cancer
and lymphoma. The RainDance platform
demonstrated the ability to quantitatively
analyze DNA methylation with high:
• Completeness (coverage at 1x of 97.3%;
average depth of coverage 313x)
• Specificity (76.8% of mapped reads
were on target)
• Uniformity (base coverage at 0.2x;
75.9% average depth of coverage)
Overview
Recent advances in high-throughput sequencing technologies have enabled the analysis of methylation patterns on a genome-wide scale, which has led to the discovery of epigenetic variations associated with cell differentiation and with the progression of diseases such as cancer. Of particular
interest are the methylation patterns from CpG islands and their role in gene regulation. Previous
techniques, including ChIP-chip, ChIP-seq, MEDiP, MSRE-seq, and whole-genome sequencing of DNA
treated with sodium bisulfite, fail to provide a complete picture of methylation activity across these
regions. RainDance Technologies has extended the utility of its targeted sequencing platform,
enabling researchers to overcome the hurdles associated with traditional techniques and accurately
target genomic regions of methylation.
Power of the RainDance Platform
RainDance Technologies has expanded the capabilities of its sophisticated primer design pipeline to
enable analysis of methylated regions of the human genome using targeted bisulfite sequencing.
Used in conjunction with high-throughput DNA sequencing, the RainDance Methyl-Seq™ capabilities
provide methylation status of specific regions across the genome with the same complete sequence
coverage, specificity, and uniformity as the company’s current Sequence Enrichment Solution.
DNA Methylation Analysis of Artificially Methylated Test Samples — The Scripps Research Institute
A primer library designed to amplify 642 loci-containing regions of known differentially methylated
CpG islands was used to sequence-enrich two genomic DNA samples. One of the two samples consisted of wild-type lymphoblast genomic DNA (red — 100% mix) and the second sample consisted of
a mixture of 40% wild-type lymphoblast genomic DNA and 60% lymphoblast genomic DNA that had
been methylated (blue — 60% mix). The resulting PCR products were sequenced using an Illumina
GAII. The diagram below shows one example in which differential methylation patterns are observed
in two adjacent genes.
Differential Methylation Patterns in the SGCE and PEG10 Genes
SGCE
PEG10
Exons
Amplicons
94122000
80
% Methylation
RDT Platform Advantages
• Up to 4,000 customer-defined regions
of interest can be interrogated at the
same time
• Sophisticated primer design algorithm
capable of interrogating all regions of
the methylome
• Ability to measure multiple cytosine
residues in a single CpG island on the
same DNA strand
• Single-template PCR eliminates competitive amplification bias due to differentially methylated target sequences
• Fully automated and easy-to-use
instrumentation
• More than 1 million PCR reactions per
sample
• Workflow complementary to all current and future sequencing platforms
• Cost-effective sequencing and simplified data analysis compared to
genome-wide approach
94122500
94123000
94123500
94124000
94124500
94125000
80
70
70
60
60
50
50
40
40
30
30
20
20
10
10
0
0
Sequencing Reads
Source: The Scripps Research Institute and RainDance Technologies
RainStorm
The RainStorm Difference
The RainStorm™ microdroplet-based technology utilized in the RDT 1000 instrument goes far beyond
conventional microfluidics, delivering on the promise of efficient, effective molecular biology at small
scale and high speed. The following performance characteristics provide gold-standard results:
Consistency. RainStorm results demonstrate less than 1% variation in droplet size. This allows the
development of statistically powerful assays with reproducible experimental conditions, robust
operation, and simplified procedures.
Stability. Exclusive innovations in oil and surfactant chemistry produce droplets that are extremely
stable. This permits shipment and storage for extended periods — without the loss of droplet integrity.
Control. Innovative design and near-real-time signal processing furnish the fine control needed to
manipulate the droplets at high rates (3,000 events/second).
RDT 1000 Specifications
RainDance Technologies, Inc.
44 Hartwell Avenue
Lexington, MA 02421, U.S.A.
Phone: 781-861-6300
Phone, toll-free: 888-724-6440
Fax: 781-861-1233
www.raindancetech.com
E-mail: [email protected]
Width
24.86 in
Depth
27.36 in
Height
19.04 in
Weight
150 lb
Power source
100/240 V AC, 50/60 Hz
Operating environment
15 – 30° C
Thermal control
Samples maintained at 10° C
Droplet volume
8 pL – 18 pL
Droplet throughput
Up to 3000 droplets per second
Service Providers
RainDance Technologies maintains a global service provider network, including:
Expression Analysis
www.expressionanalysis.com
Ambry Genetics
Atlas Biolabs
www.ambrygen.com
www.atlas-biolabs.de
Bioinformatics Partners
RainDance Technologies also partners with leading data analysis companies, such as:
RainDance and RainStorm are trademarks
of RainDance Technologies, Inc. All other
brands may be trademarks of their
respective holders.
© 2011 RainDance Technologies, Inc. All
rights reserved. Printed in U.S.A. Pub
1070045 04/11
LCN: 50-02287 Rev. B
CLCbio
www.clcbio.com