SOLiD SYSTEM

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


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.


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.

C.

D.

A.

LIB

RA

RY

PR

EPA

RAT

ION

E.

DU

AL IN

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F EA

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APPLIED BIOSYSTEMS | ThE FuTurE IS SOLiD 3


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)




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.


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.


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


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.


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.


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


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


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


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


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.


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.


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


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



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.


SOLiD SYSTEM

Documents

primer number

new primer

primer complementary

primer reset1

primer resete

this4universal seq primer

solid foundation

the5 universal seq primer