Specification Sheet: Sequencing

cBot
Fully automated clonal cluster generation for Illumina sequencing.
Illumina cBot Highlights

Figure 1: Cluster Generation is Streamlined
and Efficient on cBot

• Fast, Efficient Workflow:
Amplify > 96 samples in ~4–5 hours with < 10 minutes ofhands-on time
Library
Preparation• Easiest to Use:
Pre-packaged 96-well TruSeq™ reagents, and simple touch
screen interface simplifies operation

Sequencing• Innovative System Design:
Real-time fluidic monitoring, integrated system sensors andremote monitoring ensure robust instrument operation• Highest Quality Results:
Improved chemistry generates higher density clusters andsequencing accuracy

Cluster Generation
on cBot

Hands-on Time1

The Best Next-Gen Sequencing Workflow Just
Got Better
cBot is a revolutionary automated clonal amplification system at thecore of the Illumina sequencing workflow (Figure 1, upper panel). cBot
replaces a lab full of equipment with a single compact device, delivering unsurpassed efficiency and ease of use for the highest quality
sequencing results.
With cBot, hands-on time is reduced to less than 10 minutes, compared to more than six hours of hands-on effort for emulsion PCR
methods. The process of creating sequencing templates is complete
in about four hours, compared to more than 24 hours for emulsion
PCR-based protocols (Figure 1, lower panel).

Breakthrough System for Cluster Generation
The Illumina sequencing workflow is based on three simple steps:
libraries are prepared from virtually any nucleic acid sample, amplified
to produce clonal clusters, and sequenced using massively parallel
synthesis. The cBot clonal amplification system has innovative featuresthat eliminate user intervention, reduce potential failure points, andincrease sequencing productivity.
TruSeq Cluster Generation reagents are packaged in ready-to-use
96-well plates, completely removing reagent preparation errors,potential sources of contamination, and decreasing storage requirements. cBot features a single unique, plate-piercing manifold forintervention-free runs. Cluster generation occurs within the sealed,eight-channel Illumina flow cell, bypassing the frequent handling andcontamination issues inherent to emulsion PCR-based protocols.
cBot is capable of processing > 96 samples within a single flow cell,
resulting in substantial cost savings without incremental effort andwasted reagents. Innovative instrument features ensure seamless
operation for your sequencing workflow (Figure 2).

3

Total Process Time* (Hours)(Hours)66

Illumina
cBot

Illumina
cBot

Illumina
Cluster
StationIllumina
Cluster
StationEmulsion
PCR

Emulsion
PCR

1224*Unlike emulsion PCR methods, processing multiplexed runs (>96 samples) using cBot requires no additional time

cBot performs automated simultaneous clonal amplification of hundreds ofmillions of single-molecule DNA templates, dramatically reducing hands-on
and total preparation time required to produce sequencing template, compared to emulsion PCR methods.

Better Results with Less Effort
cBot software enhancements and user interaction features ensure highproductivity:• Integrated 8-inch touch screen provides simplified operation ina small, lab-friendly footprint• On-screen, step-by-step instructions with embedded
multimedia help enable user operation with no prior training• Real-time progress indicators provide at-a-glance monitoring• Remote monitoring allows a single user to manage multiple
systems from any web browser or phone• Status emails are sent when the run is complete or whenintervention is required

cBot Cluster Generation Process
Prior to sequencing, single-molecule DNA templates are bridge
amplified to form clonal clusters inside the flow cell. (Figure 3).

Specification Sheet: Sequencing
Figure 2: New cBot Features Enables Rapid and Streamlined Cluster Generation
Single disposable
flow cell manifoldwith self-piercing
sippers eliminatesuser intervention

Thermal stage enables
end-to-end temperature
uniformity for optimalcluster density

Manifold clamps forleak-free connections
and superior thermal
contact
Eight-channel flow cell
reduces risk of contaminationand eliminates the needfor extra equipment

Touch screen monitor
simplifies operationand provides real-time
run monitoring

Ready-to-use 96-wellreagent plate eliminates
preparation errors

Easy-access waste
bottle chamber withliquid level sensor

Integrated barcode
reader for automatic
information capture

The cBot cluster generation system is the next generation of workflow improvements for Illumina sequencing. Novel innovations
include pre-packaged reagents, a single manifold, advanced fluidics and thermal stage features, integrated sensors, remotemonitoring capabilities, and simplified data entry and tracking with the touch screen and barcode scanner.

Specification Sheet: Sequencing
Figure 3: Cluster Generation by Isothermal Bridge Amplification

Cluster generation from single-molecule DNA templates occurs within the sealed Illumina flow cell on the cBot instrument, and involves immobilization and 3’ extension, bridge amplification, linearization, and hybridization.

Immobilization of Single-Molecule DNA
Templates

Linearization, Blocking, and PrimerHybridization

Hundreds of millions of templates are hybridized to a lawn of oligonucleotides immobilized on the flow cell surface. The templates arecopied from the hybridized primers by 3’ extension using a high-fidelity
DNA polymerase to prevent misincorporation errors. The original
templates are denatured, leaving the copies immobilized on the flow
cell surface.

Each cluster of dsDNA bridges is denatured, and the reverse strand isremoved by specific base cleavage, leaving the forward DNA strand.
The 3’-ends of the DNA strands and flow cell-bound oligonucleotides
are blocked to prevent interference with the sequencing reaction. The
sequencing primer is hybridized to the complementary sequence on
the Illumina adapter on unbound ends of the templates in the clusters.
The flow cell now contains >200 million clusters with ~1,000 molecules/cluster, and is ready for sequencing.

Isothermal Bridge Amplification
Immobilized DNA template copies are amplified by isothermal bridge
amplification. The templates loop over to hybridize to adjacent lawn
oligonucleotides. DNA polymerase copies the templates from thehybridized oligonucleotides, forming dsDNA bridges, which are denatured to form two ssDNA strands. These two strands loop over andhybridize to adjacent oligonucleotides and are extended again to form
two new dsDNA loops. The process is repeated on each template bycycles of isothermal denaturation and amplification to create millions ofindividual, dense clonal clusters containing ~2,000 molecules.

Summary
Illumina sequencing with cBot automated cluster generation sets thenew standard for simplified next- generation sequencing. Ready-touse reagents, smart instrumentation improvements, and new cluster
generation chemistry offers significant advantages over emulsion
PCR-based workflows and promotes even higher data density andsequencing accuracy. By streamlining the critical clonal amplification
step in the next-generation sequencing workflow, Illumina continues toaccelerate your landmark discoveries and publications.

Ordering Information
Description

Catalog No.
cBotHiSeq System

Genome Analyzer

TruSeq Single-Read Cluster Generation Kits include flow cell,
reagent plate, manifold, user instructions

GD-401-3001

GD-300-2001

TruSeq Paired-End Cluster Generation Kits include flow cell,reagent plate, manifold, PE reagents, user instructions

PE-401-3001

PE-300-2001

cBot Instrument Includes cBot, flow cell adapter plate,
one year warranty, user manual

SY-301-2002

cBot Flow Cell Manifold (Optional)

SY-301-2014

Specification Sheet: Sequencing
cBot System Specifications
Catalog No.
SY-301-2002
Instrument Configuration
CE Marked and ETL Listed instrument, Installation, setup, andaccessories
Instrument Control Computer
Mini-ITX Board with Celeron M Processor

ADDITIONAL INFORMATION
For more information about
Illumina sequencing, visit www.illumina.com/sequencing or contact us
at the address below.

Laser radiation
Do not stare into the visible-light beam of the barcode scanner.
The barcode scanner is a Class 2 laser product.

1 GB RAM, 80 GB Hard Drive
Windows Embedded OS
Integrated 8” Touch Screen Monitor
Operating Environment
Temperature: 22°C ± 3°C
Humidity: Non-Condensing 20%–80%
Altitude: Less than 2,000 m (6,500 ft)
Air Quality: Pollution Degree Rating of II
For Indoor Use OnlyLaser
Class 2 Laser: 630­–650 nm
Dimensions
W×D×H: 38 cm × 62 cm × 40 cmWeight: 34 kg
Crated Weight: 36 kg
Power Requirements
100−240V AC 50/60 Hz, 4A, 400 Watts

Illumina, Inc. • 9885 Towne Centre Drive, San Diego, CA 92121 USA • 1.800.809.4566 toll-free • 1.858.202.4566 tel • techsupport@illumina.com • illumina.com
For research use only
© 2011 Illumina, Inc. All rights reserved.
Illumina, illuminaDx, BeadArray, BeadXpress, cBot, CSPro, DASL, Eco, Genetic Energy, GAIIx, Genome Analyzer, GenomeStudio,GoldenGate, HiScan, HiSeq, Infinium, iSelect, MiSeq, Nextera, Sentrix, Solexa, TruSeq, VeraCode, the pumpkin orange color, and
the Genetic Energy streaming bases design are trademarks or registered trademarks of Illumina, Inc. All other brands and names
contained herein are the property of their respective owners.
Pub. No. 770-2009-032 Current as of 27 April 2011