Next Generation Sequencing with SOLiD System

1. The SOLiD 3 System ™
Enabling the Next Generation of Science

2. Revolutionary Technology
SOLID Platform
Applied Biosystems has been the leader in sequence Imagine the scientific discoveries that can be achieved with the power of the SOLiD 3 System:
analysis for more than 20 years and has revolutionized the
industry by developing innovative chemistries and robust, • The power to view the whole transcriptome — without any holes.
automated instrumentation. The SOLiD™ 3 System is • The ability to distinguish strand specific expression patterns — for the first time.
Applied Biosystems latest revolutionary step forward in • The accuracy to detect SNPs at low coverage — with a low false positive rate.
sequence analysis technology. The throughput capability • The capacity to globally assess DNA-protein binding interactions — across multiple samples
and unrivaled accuracy inherent in the SOLiD 3 System, in a single run.
coupled with broad application flexibility, provide a unique • The sensitivity to discover novel transcripts and splice variation — without the bias of microarrays.
system for advancing your research like never before. • The resolution to characterize structural rearrangements — including balanced translocations.

3. Compute cluster for real-time data analysis SOLiD 3 System Side and bottom reagent storage for walk away operations
Pre-filled reagent strip tubes Dual independent flow cells
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4. SOLiD™ System Chemistry | SOLID INSIDE AND OUT
A. LIBrArY PrEPArATIOn
The SOLiD 3 System uses two types of
libraries—fragment or mate-paired. The type
LIBRARY PREPARATION
of library used depends on the application and
information required.
A.
B. EMuLSIOn PCr/BEAD EnrIChMEnT
Clonal bead populations are prepared in
microreactors containing template, PCr
reaction components, beads and primers.
After PCr, the templates are denatured and
bead enrichment is performed to select beads
with extended templates. The template on the
selected beads undergoes a 3’ modification to
allow covalent bonding to the slide.
B.
C. BEAD DEPOSITIOn
3’ modified beads are deposited onto a glass
slide. Deposition chambers offer the ability
to segment a slide into one, four or eight
chambers during the bead loading process. A
key advantage of the system is the ability to
accommodate increasing densities of beads
per slide, which will result in a higher level of
throughput from the same system.
C.
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5. P1 Adapter TA Template Sequence TA
2. Image 5. Repeat steps 1-4 to Extend Sequence
Excite Fluorescence
Ligation cycle 1 2 3 4 5 6 7 ... (n cycles)
AT TT CT GT TT CA GC D. SEquEnCInG BY LIGATIOn
3’ TA AA GA CA AA GT CG
D.
TA 3’
Primers hybridize to the P1 adaptor
sequence within the library template. A set
of four fluorescently labeled di-base probes
compete for ligation to the sequencing
3. Cap Unextended Strands 6. Primer Reset primer. Specificity of the di-base probe
is achieved by interrogating every 1st
Phosphatase and 2nd base in each ligation reaction.
PO4 Universal seq primer (n-1) Multiple cycles of ligation, detection and
3’ 2. Primer reset 1. Melt off extended
cleavage are performed with the number
sequence
1 m
of cycles determining the eventual read
3’ bead 3’
length. Following a series of ligation
cycles, the extension product is removed
and the template is reset with a primer
complementary to the n-1 position for a
7. Repeat steps 1-5 with new primer second round of ligation cycles.
1 base shift
PRIMER ROUND 2
-1
Universal seq primer (n-1) AA CA CG TC AA TA CC
3’
1 m
bead 3’
T GT GC AG TT AT GG
E. PrIMEr rESET
E.
8. Repeat Reset with , n-2, n-3, n-4 primers
Five rounds of primer reset are completed
Read Position 0 1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 for each sequence tag. Through the primer
DUAL INTERROGATION OF EACH BASE
Universal seq primer (n) reset process, each base is interrogated
1
3’ in two independent ligation reactions
Universal seq primer (n-1) by two different primers. For example,
2 3’
Primer Round
the base at read position 5 is assayed by
3 Universal seq primer (n-2) primer number 2 in ligation cycle 2 and by
3’
primer number 3 in ligation cycle 1. This
4 Universal seq primer (n-3) dual interrogation is fundamental to the
3’
unmatched accuracy characterized by the
5 Universal seq primer (n-4)
3’ SOLiD System.
Indicates positions of interrogation Ligation Cycle 1 2 3 4 5 6 7
To view an animation of the SOLiD System,
visit solid.appliedbiosystems.com.
APPLIED BIOSYSTEMS | ThE FuTurE IS SOLiD 3
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6. Your Science Deserves
a SOLID Foundation
The SOLiD™ 3 System is revolutionary in terms of throughput, and differentiates itself from
other next-generation platforms with unparalleled accuracy, system robustness and scalability.
Walk away automation and enhanced data analysis tools significantly reduce hands on
time and expedite your time to results. With longer read lengths and sample multiplexing
capabilities, the SOLiD 3 System provides the most cost effective analysis for a wide range
of genomic, transcriptomic and epigenomic applications.
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7. System Scalability Superior Accuracy
The SOLiD 3 System’s open slide format and flexible bead densities With system accuracy greater than 99.94%, due to 2 base encoding,
enable increases in throughput with modest analysis and chemistry the SOLiD 3 System distinguishes itself by providing data that is
optimizations. While competitive technologies are already near more accurate than alternative next-generation platforms for variation
maximum throughput, detection. This unrivaled accuracy is inherent in the system and results
the SOLiD 3 System is in lower false
Superior Accuracy
scalable to meet your positive rates.
research needs today 40 40%
35 35%
and tomorrow.
MEASURED QUALITY VALUE
30 30%
25%
ERROR RATE
25
20 20%
15 15%
10 10%
0 0%
0 5 10 15 20 25 30 35 40
POSITION IN READ BP
Unmatched Throughput Maximum Flexibility
The SOLiD 3 System can generate over 20 gigabases of mappable data The independent flow cells of the SOLiD 3 System enable users to run two
and more than 400 million tags per run, which is more usable data than completely independent experiments on one SOLiD Analyzer – providing two
any other next-generation instruments in one.
system available Sample multiplexing Unsegmented
Quad Octet
Full Slide
today. This throughput 50 with barcoding
translates into 45 sequences provides
AB R&D Individual
efficient sequencing 40
CUSTOMER LAB
additional flexibility Samples
of multiple genomes 35 and dramatically 2 samples/run 8 samples/run 16 samples/run
or cost effective 30 increases the cost-
GB/RUN
analysis of multiple 25
effectiveness of Multiplexed
transcriptomes. 20
targeted resequencing, Samples (20
barcodes)
15
expression and ChIP ≤20 samples/slide
≤20 samples/segment
≤4 segments/slide
≤20 samples/segment
≤8 segments/slide
10
analyses. x 2 slides/run
≤ 40 samples/run
x 2 slides/run
≤ 160 samples/run
x 2 slides/run
≤ 320 samples/run
0
OCTOBER ‘07 FEBRUARY ‘08 MAY ‘08 OCTOBER ‘08 FEBRUARY ‘09
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8. SOLID Science
SOLID Applications
Increase your research productivity with the maximum flexibility, superior accuracy and unmatched throughput of the
SOLiD™ 3 System. Walk away automation and enhanced data analysis tools significantly reduce hands on time and
expedite your time to results. With longer read lengths and sample multiplexing capabilities, the SOLiD 3 System provides
the most cost effective analysis for a wide range of genomic, transcriptomic and epigenomic applications.
Whole Transcriptome Analysis Whole Genome Resequencing Targeted Resequencing
The SOLiD 3 System provides researchers The SOLiD 3 System enables a new level of whole The superior accuracy of the SOLiD 3 System
with access to the entire transcriptome with a genome sequencing with massive throughput and provides a very low false positive rate that enables
hypothesis generating, highly sensitive assay mate-paired technology. The system’s ultra high variation detection with less coverage than
that preserves the strand information of the throughput results in high coverage levels and the alternative platforms. The inclusion of barcodes
original RNA molecule. The combination of 400 M ability to sequence multiple genomes in a single for sample multiplexing dramatically lowers the
sequence tags and sample multiplexing capabilities run. The combination of unmatched throughput and cost, time and labor involved in processing
allow cost effective analysis of multiple samples superior accuracy dramatically reduces the time multiple samples.
in a single run while maintaining the sensitivity to and cost to lower than that of other technologies.
discover molecules present at low levels.
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9. Complete solutions for your experimental workflow.
DESIGN ISOLATE PREPARE LIBRARY AMPLIFY & ANALYZE DATA VALIDATE
EXPERIMENT
N NUCLEIC ACID SEQUENCE RESULTS
20
10
5
0
0 5 10 20
0
0 5
5 01
01 02
02
0
5
01
02
TNEMIREPXE
N DICA CIELCUN ECNEUQES STLUSER
NGISED YRARBIL ERAPERP ATAD EZYLANA
ETALOSI & YFILPMA ETADILAV
Small RNA Analysis into a scaffold that can then be filled in with the SOLiD System’s flexible slide format and
The SOLiD Small RNA Solution is a robust method capillary electrophoresis-based sequencing. multiplexing capability allows researchers to
for hypothesis-generating, whole genome analysis analyze multiple samples in a single run.
of expression patterns that enable discovery of Gene Expression Profiling
novel RNA without the probe bias of microarrays. The massive amount of sequence tags generated Methylation Analysis
Using the SOLiD Small RNA Expression Kit, you in a single run can be used to identify and quantify The SOLiD 3 System provides a high throughput
can convert total RNA into a library suitable for exon sequences associated with alternative method for analyzing genome-wide DNA
emulsion PCR using five easy steps with a single splicing and promoter usage. Simple tag counting methylation with a much higher level of resolution
purification. This simplified protocol conserves allows for digital readout of expression levels and than traditional methods.
the strand information of the original molecule the ability to detect lowly expressed genes with
and enables one to decipher strand-specific greater sensitivity.
expression patterns.
Chromatin Immunoprecipitation (ChIP)
ChIP-Sequencing (ChIP-Seq) with the SOLiD 3
De Novo Sequencing
System combines a chromatin immunoprecipitation
The longer read lengths and new data analysis
(ChIP) assay with massively parallel DNA
software of the SOLiD 3 System enable the de
sequencing, offering a highly sensitive method
novo assembly of small organisms. Researchers
to accurately characterize the protein-DNA
have demonstrated the assembly of 50 bp reads
interactions of an entire genome. Additionally,
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10. SOLID Software. Your Analysis Solution.
Large scale science requires solid analysis SOLID Data Acquisition SOLID Analysis SOLID Interpretation
The SOLiD Instrument Control The SOLiD Analysis Tools The SOLiD Alignment Browser
solutions. That’s why the SOLiD 3 System
™ Software (ICS) performs image (SAT) provide a powerful back (SAB) is an open-source
acquisition and quality control. end engine for automated data platform to enable visualization
is designed to deliver not only the computing The ICS provides an easy-to-use analysis that generates base space of sequences aligned to
interface for walk away automation. results. The SOLiD Experimental a reference genome. This
power, but the software solutions to support Tracking System (SETS) is a allows further visualization and
web-based integrated application discrimination of experimental
all of your next generation projects. From image for real-time remote monitoring results without the need for
and report viewing. A custom additional computing power.
acquisition to biological interpretation, the analysis pipeline can be integrated
with SAT/SETS via SOLiD
integration of the SOLiD Software Suite and
Applications Interface (SAI).
tools from the SOLiD Software Community
support a broad range of application-specific
analyses for your genomic, epigenomic or
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
transcriptomic research.
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
ACGT ACGT ACGT ACGT
Data Acquisition Sequence Analysis Biological Interpretation
Application-Specific Analyses
SOLID Applications
The SOLiD Applications Interface integrates
application-specific tools from the SOLiD Software
Community with the Applied Biosystems’ Software
Suite for seamless analysis of your data.
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11. Join the SOLiD Software Community
The SOLiD Software Community provides a resource to those seeking application-specific tools or in
developing new tools for analysis of SOLiD System data. The goal of this program is to directly address
the challenges associated with analyzing and managing the vast amounts of research data generated
by this ultra high throughput sequencing technology. Resources available through the program provide
SOLID researchers with solutions for all supported
applications including whole genome and targeted
resequencing, whole transcriptome, de novo sequencing,
ChIP-Seq and methylation studies.
Academic AB Open
Partner
Open Source Source Analysis
Software
Tools Tools
Visit the community today at
info.appliedbiosystems.com/solidsoftwarecommunity
PUBLICATION
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12. The Future Is SOLID
The ability to increase both data output and sample throughput will be a key driver for
pushing the boundaries of biological analysis in the years to come. The SOLiD™ 3 System’s
open slide format is amenable to vastly increasing bead densities and increasing read length
without sacrificing robustness or accuracy. Increases in bead density come with minor
incremental costs and will drive significant reductions in the overall cost per run. These
features will ensure that today’s system will
meet the needs of future experiments by growing
and expanding with your research.
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13. Increasing Future Data Output and Throughput Bead Density — Amount of beads that can be deposited on a
slide and analyzed.
The SOLiD 3 System’s ability to dramatically increase data output The unique open format of the SOLiD 3 System allows more beads to
using the same platform is a function of the flexible and modular be deposited on a single slide, than with microtiter-based technologies,
nature of the technology. Achieving higher levels of increased data without changing the system configuration. Advancements in the
output is possible with modest improvements. algorithms for bead finding will further increase the number of beads that
can be analyzed from a given slide. These combined improvements are
anticipated to more than double the data output of the current system.
50
45
AB R&D
40
CUSTOMER LAB
35
30
GB/RUN
25
20
15
10
0
OCTOBER ‘07 FEBRUARY ‘08 MAY ‘08 OCTOBER ‘08 FEBRUARY ‘09
read Length— Measure of the number FrAGMEnT LIBrArY MATE-PAIrED LIBrArY
of bases sequenced. using two full slides/run using two full slides/run
Protocol and chemistry optimization will 2007 2008 2009 2010 2007 2008 2009 2010
increase the read length of the current read Length 35 35 50 100 25 35 50 100
system, while maintaining strict accuracy Total Output* ~2.5 GB ~4 GB ~10 GB >35 GB ~3.5 GB ~12 GB >20 GB >50 GB
and consistency standards.
* Mappable data from 2 slide run
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14. Your Success Depends on SOLID Support
Global Service & Support
With more than 2,000 field personnel, Applied Biosystems has the most
extensive network of dedicated Field Application Specialists (FAS) and
Support Engineers to work with you. With Applied Biosystems, you can feel
confident that we will be there when and where you need us most.
World Class Training Programs*
Regionally located training facilities offer courses in Library prep,
System Sequencing and Bioinformatics.
Europe:
330 Field Sales
270 Field Service Asia Pacific:
*
Americas: 150 Supply Chain * 200 Field Sales
Headquarters: 350 Field Sales
H 200 Field Service
120 Direct * 300 Field Service
40 Supply Chain
Global Support 210 Supply Chain
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16. Applied Biosystems
850 Lincoln Centre Drive
Foster City, CA 94404
USA
For Research Use Only. Not for use in diagnostic procedures. © 2009 Applied Biosystems. Applera, Applied Biosystems, and AB (Design) are registered trademarks and SOLiD
is a Research UseApplera Corporationdiagnostic procedures. US and/or certain other countries.
For trademark of Only. Not for use in or its subsidiaries in the
All2009 Applied Biosystems.sole property of their and AB (Design) are01/2009 trademarks and SOLiD is a trademark of Applied Biosystems or its subsidiaries
© other trademarks are the Applied Biosystems, respective owners. registered
in the US and/or certain other countries. All other trademarks are the sole property of their respective owners.
Publication 106BR13-01
Publication 139BR01-03