solid system
DESCRIPTION
Next Generation Sequencing with SOLiD SystemTRANSCRIPT
Enabling the Next Generation of ScienceThe SOLiD™ 3 System
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 trademark of Applera Corporation or its subsidiaries in the US and/or certain other countries. All other trademarks are the sole property of their respective owners. 01/2009
Publication 106BR13-01
Applied Biosystems 850 Lincoln Centre Drive Foster City, CA 94404 USA
For Research Use Only. Not for use in diagnostic procedures. © 2009 Applied Biosystems. Applied Biosystems, and AB (Design) are registered trademarks and SOLiD is a trademark of Applied Biosystems or its subsidiaries in the US and/or certain other countries. All other trademarks are the sole property of their respective owners.
Publication 139BR01-03
Revolutionary Technology SOLID Platform
Applied Biosystems has been the leader in sequence analysis for more than 20 years and has revolutionized the industry by developing innovative chemistries and robust, automated instrumentation. The SOLiD™ 3 System is Applied Biosystems latest revolutionary step forward in sequence analysis technology. The throughput capability and unrivaled accuracy inherent in the SOLiD 3 System, coupled with broad application flexibility, provide a unique system for advancing your research like never before.
Imagine the scientific discoveries that can be achieved with the power of the SOLiD 3 System: • The power to view the whole transcriptome — without any holes. • The ability to distinguish strand specific expression patterns — for the first time. • The accuracy to detect SNPs at low coverage — with a low false positive rate. • The capacity to globally assess DNA-protein binding interactions — across multiple samples in a single run. • The sensitivity to discover novel transcripts and splice variation — without the bias of microarrays. • The resolution to characterize structural rearrangements — including balanced translocations.
APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS 2
Pre-filled reagent strip tubes
SOLiD 3 System Side and bottom reagent storage for walk away operationsCompute cluster for real-time data analysis
Dual independent flow cells
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 of library used depends on the application and information required. 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.
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.
B.
C.
D.
A.
LIB
RA
RY
PR
EPA
RAT
ION
E.
DU
AL IN
TER
RO
GATIO
N O
F EA
CH
BA
SE
3 APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS
D. SEquEnCInG BY LIGATIOn 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 primer. Specificity of the di-base probe is achieved by interrogating every 1st and 2nd base in each ligation reaction. Multiple cycles of ligation, detection and cleavage are performed with the number of cycles determining the eventual read 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 second round of ligation cycles.
B.
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A.
LIB
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ION
E.
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APPLIED BIOSYSTEMS | ThE FuTurE IS SOLiD 3
APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS 4
Ligation Cycle
Prim
er R
ound
1 2 3 4 5 6 7
3’AT
TA
Universal seq primer (n)
3’P1 Adapter Template Sequence
POH
Universal seq primer (n-1)
Ligase
Phosphatase
+
1. Prime and Ligate
2. Image 5. Repeat steps 1-4 to Extend Sequence
3’
Universal seq primer (n-1)1. Melt off extended sequence
2. Primer reset3’
AA CA
GCGT
CG TC AA
AG GG
CC
T TT
6. Primer Reset
7. Repeat steps 1-5 with new primer
8. Repeat Reset with , n-2, n-3, n-4 primers
AT
TA
TA3’
Excite Fluorescence
ATAA GA CA AATATT CT GT TT CA
GTGCCG
3’
4. Cleave off Fluor
AT
TA3’
Cleavage Agent
P
HO
3. Cap Unextended Strands
3’
PO4
1 2 3 4 5 6 7 ... (n cycles)Ligation cycle
3’
3’1 μmbead
1 μmbead
1 μmbead
-1
Universal seq primer (n-1)
Universal seq primer (n)
Universal seq primer (n-2)
Universal seq primer (n-3)
Universal seq primer (n-4)
3’
3’
3’
3’
3’
1
2
3
4
5
PRIMER ROUND 1
PRIMER ROUND 2
1 base shift
Read Position
Indicates positions of interrogation
35343332313029282726252423222120191817161514131211109876543210
To view an animation of the SOLiD System, visit solid.appliedbiosystems.com.
E. PrIMEr rESET Five rounds of primer reset are completed for each sequence tag. Through the primer reset process, each base is interrogated in two independent ligation reactions by two different primers. For example, the base at read position 5 is assayed by primer number 2 in ligation cycle 2 and by primer number 3 in ligation cycle 1. This dual interrogation is fundamental to the unmatched accuracy characterized by the SOLiD System.
Your Science Deserves a SOLID FoundationThe 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.
5 APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS
System Scalability
The SOLiD 3 System’s open slide format and flexible bead densities enable increases in throughput with modest analysis and chemistry optimizations. While competitive technologies are already near maximum throughput, the SOLiD 3 System is scalable to meet your research needs today and tomorrow.
APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS 6
Maximum Flexibility
The independent flow cells of the SOLiD 3 System enable users to run two completely independent experiments on one SOLiD Analyzer – providing two instruments in one. Sample multiplexing with barcoding sequences provides additional flexibility and dramatically increases the cost- effectiveness of targeted resequencing, expression and ChIP analyses.
Superior Accuracy
With system accuracy greater than 99.94%, due to 2 base encoding, the SOLiD 3 System distinguishes itself by providing data that is more accurate than alternative next-generation platforms for variation detection. This unrivaled accuracy is inherent in the system and results in lower false positive rates.
2 samples/run 8 samples/run 16 samples/run
≤20 samples/slidex 2 slides/run
≤ 40 samples/run
≤20 samples/segment≤4 segments/slide
x 2 slides/run
≤ 160 samples/run
≤20 samples/segment≤8 segments/slide
x 2 slides/run
≤ 320 samples/run
Individual Samples
Multiplexed Samples (20 barcodes)
Unsegmented Full Slide
Quad
Octet
ERROR RATE
40
35
30
25
20
15
10
0
Superior Accuracy
MEA
SURE
D QU
ALIT
Y VA
LUE
40%
35%
30%
25%
20%
15%
10%
0%
0 105 15 20 25 30 35 40
POSITION IN READ BP
Unmatched Throughput
The SOLiD 3 System can generate over 20 gigabases of mappable data and more than 400 million tags per run, which is more usable data than any other next-generation system available today. This throughput translates into efficient sequencing of multiple genomes or cost effective analysis of multiple transcriptomes.
13.4 GB/run
17 GB/run
60
40
50
30
20
10
AB R&DCustomer Lab
OCTOBER ‘07
50
45
40
35
30
25
20
15
10
0
FEBRUARY ‘08 MAY ‘08 OCTOBER ‘08 FEBRUARY ‘09
GB/R
UN
AB R&D
CUSTOMER LAB
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.
SOLID Science SOLID Applications
7 APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS
Whole Transcriptome Analysis The SOLiD 3 System provides researchers with access to the entire transcriptome with a hypothesis generating, highly sensitive assay that preserves the strand information of the original RNA molecule. The combination of 400 M sequence tags and sample multiplexing capabilities allow cost effective analysis of multiple samples in a single run while maintaining the sensitivity to discover molecules present at low levels.
Whole Genome Resequencing The SOLiD 3 System enables a new level of whole genome sequencing with massive throughput and mate-paired technology. The system’s ultra high throughput results in high coverage levels and the ability to sequence multiple genomes in a single run. The combination of unmatched throughput and superior accuracy dramatically reduces the time and cost to lower than that of other technologies.
Targeted Resequencing The superior accuracy of the SOLiD 3 System provides a very low false positive rate that enables variation detection with less coverage than alternative platforms. The inclusion of barcodes for sample multiplexing dramatically lowers the cost, time and labor involved in processing multiple samples.
Small RNA Analysis The SOLiD Small RNA Solution is a robust method for hypothesis-generating, whole genome analysis of expression patterns that enable discovery of novel RNA without the probe bias of microarrays. Using the SOLiD Small RNA Expression Kit, you can convert total RNA into a library suitable for emulsion PCR using five easy steps with a single purification. This simplified protocol conserves the strand information of the original molecule and enables one to decipher strand-specific expression patterns. De Novo Sequencing The longer read lengths and new data analysis software of the SOLiD 3 System enable the de novo assembly of small organisms. Researchers have demonstrated the assembly of 50 bp reads
into a scaffold that can then be filled in with capillary electrophoresis-based sequencing.
Gene Expression Profiling The massive amount of sequence tags generated in a single run can be used to identify and quantify exon sequences associated with alternative splicing and promoter usage. Simple tag counting allows for digital readout of expression levels and the ability to detect lowly expressed genes with greater sensitivity.
Chromatin Immunoprecipitation (ChIP) ChIP-Sequencing (ChIP-Seq) with the SOLiD 3 System combines a chromatin immunoprecipitation (ChIP) assay with massively parallel DNA sequencing, offering a highly sensitive method to accurately characterize the protein-DNA interactions of an entire genome. Additionally,
the SOLiD System’s flexible slide format and multiplexing capability allows researchers to analyze multiple samples in a single run.
Methylation Analysis The SOLiD 3 System provides a high throughput method for analyzing genome-wide DNA methylation with a much higher level of resolution than traditional methods.
APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS 8
ISOLATE NUCLEIC ACID
PREPARE LIBRARY AMPLIFY & SEQUENCE
ANALYZE DATA VALIDATE RESULTS
20
0
5
10
0 5 10 20
DESIGN EXPERIMENNT
20
0
5
10
0 5 10 20
ISOLATE NUCLEIC ACID
PREPARE LIBRARY AMPLIFY & SEQUENCE
ANALYZE DATA VALIDATE RESULTS
20
0
5
10
0 5 10 20
DESIGN EXPERIMENNT
20
0
5
10
0 5 10 20
Complete solutions for your experimental workflow.
SOLID Software. Your Analysis Solution.
Large scale science requires solid analysis
solutions. That’s why the SOLiD™ 3 System
is designed to deliver not only the computing
power, but the software solutions to support
all of your next generation projects. From image
acquisition to biological interpretation, the
integration of the SOLiD Software Suite and
tools from the SOLiD Software Community
support a broad range of application-specific
analyses for your genomic, epigenomic or
transcriptomic research.
9 APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS
SOLID Interpretation
The SOLiD Alignment Browser
(SAB) is an open-source
platform to enable visualization
of sequences aligned to
a reference genome. This
allows further visualization and
discrimination of experimental
results without the need for
additional computing power.
SOLID Analysis
The SOLiD Analysis Tools
(SAT) provide a powerful back
end engine for automated data
analysis that generates base space
results. The SOLiD Experimental
Tracking System (SETS) is a
web-based integrated application
for real-time remote monitoring
and report viewing. A custom
analysis pipeline can be integrated
with SAT/SETS via SOLiD
Applications Interface (SAI).
SOLID Data Acquisition
The SOLiD Instrument Control
Software (ICS) performs image
acquisition and quality control.
The ICS provides an easy-to-use
interface for walk away automation.
Application Specific Analyses
Application-Specific Analyses
PUBLICATION
Data Acquisition Sequence Analysis Biological Interpretation
• RNA Analysis Tools • Sequence Analysis Tools • Methylation Analysis Tools • De novo Analysis Tools • ChIP Seq Analysis Tools
SOLiD Applications Interface
Data Acquisition Sequence Analysis Biological Interpretation
SOLiD Applications Interface
SOLiD InstrumentControl Software (ICS)
SOLiD ExperimentalTracking Software (SETS)
SOLiD AlignmentBrowser (SAB)
• RNA Analysis Tools • Sequence Analysis Tools• Methylation Analysis Tools • De novo Analysis Tools• ChIP Seq Analysis Tools
Tools for Success
3rd Party Software
Academic Open Source
Tools
Data Sets
AB Open Source Analysis
Tools
Data Acquisition Sequence Analysis Biological Interpretation
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.
ACGT ACGT ACGT ACGT ACGT ACGTACGT ACGT ACGT ACGTACGT ACGTACGT ACGT ACGT ACGT ACGT ACGTACGT ACGT ACGT ACGTACGT ACGT
ACGT ACGT ACGT ACGT ACGT ACGTACGT ACGT ACGT ACGTACGT ACGTACGT ACGT ACGT ACGT ACGT ACGTACGT ACGT ACGT ACGTACGT ACGT
APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS 10
SOLID Software. Your Analysis Solution.
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.
Visit the community today at info.appliedbiosystems.com/solidsoftwarecommunity
PUBLICATION
Partner Software
Academic Open Source
Tools
AB Open Source Analysis
Tools
The Future Is SOLID
11 APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS
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.
Increasing Future Data Output and ThroughputThe SOLiD 3 System’s ability to dramatically increase data output using the same platform is a function of the flexible and modular nature of the technology. Achieving higher levels of increased data output is possible with modest improvements.
APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS 12
Bead Density — Amount of beads that can be deposited on a slide and analyzed. The unique open format of the SOLiD 3 System allows more beads to be deposited on a single slide, than with microtiter-based technologies, without changing the system configuration. Advancements in the 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.
FrAGMEnT LIBrArY MATE-PAIrED LIBrArY using two full slides/run using two full slides/run
* Mappable data from 2 slide run
2007 2008 2009 2010 2007 2008 2009 2010
read Length 35 35 50 100 25 35 50 100
Total Output* ~2.5 GB ~4 GB ~10 GB >35 GB ~3.5 GB ~12 GB >20 GB >50 GB
read Length—Measure of the number of bases sequenced. Protocol and chemistry optimization will increase the read length of the current system, while maintaining strict accuracy and consistency standards.
13.4 GB/run
17 GB/run
60
40
50
30
20
10
AB R&DCustomer Lab
OCTOBER ‘07
50
45
40
35
30
25
20
15
10
0
FEBRUARY ‘08 MAY ‘08 OCTOBER ‘08 FEBRUARY ‘09
GB/R
UN
AB R&D
CUSTOMER LAB
Headquarters:
120 Direct
Global Support
Asia Pacific:
200 Field Sales
200 Field Service
40 Supply Chain
Europe:
330 Field Sales
270 Field Service
150 Supply ChainAmericas:
350 Field Sales
300 Field Service
210 Supply Chain
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.
13 APPLIED BIOSYSTEMS | ThE SOLID GEnErATIOn DELIvErS
Your Success Depends on SOLID Support
H*
* *
Revolutionary Technology SOLID Platform
Applied Biosystems has been the leader in sequence analysis for more than 20 years and has revolutionized the industry by developing innovative chemistries and robust, automated instrumentation. The SOLiD™ 3 System is Applied Biosystems latest revolutionary step forward in sequence analysis technology. The throughput capability and unrivaled accuracy inherent in the SOLiD 3 System, coupled with broad application flexibility, provide a unique system for advancing your research like never before.
Imagine the scientific discoveries that can be achieved with the power of the SOLiD 3 System: • The power to view the whole transcriptome — without any holes. • The ability to distinguish strand specific expression patterns — for the first time. • The accuracy to detect SNPs at low coverage — with a low false positive rate. • The capacity to globally assess DNA-protein binding interactions — across multiple samples in a single run. • The sensitivity to discover novel transcripts and splice variation — without the bias of microarrays. • The resolution to characterize structural rearrangements — including balanced translocations.
Enabling the Next Generation of ScienceThe SOLiD™ 3 System
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 trademark of Applera Corporation or its subsidiaries in the US and/or certain other countries. All other trademarks are the sole property of their respective owners. 01/2009
Publication 106BR13-01
Applied Biosystems 850 Lincoln Centre Drive Foster City, CA 94404 USA
For Research Use Only. Not for use in diagnostic procedures. © 2009 Applied Biosystems. Applied Biosystems, and AB (Design) are registered trademarks and SOLiD is a trademark of Applied Biosystems or its subsidiaries in the US and/or certain other countries. All other trademarks are the sole property of their respective owners.
Publication 139BR01-03