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HiSeq® Rapid Duo cBot™ Sample
Loading Kit
Reagent Prep Guide
FOR RESEARCH USE ONLY
Reagent Kit Overview
Kit Contents and Storage Requirements
Prepare Reagent Plate
Next Steps
Technical Assistance
ILLUMINA PROPRIETARY
Part # 15059382 Rev. A
Catalog # CT-403-9001DOC
November 2014
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consent of Illumina. Illumina does not convey any license under its patent, trademark, copyright, or commonlaw rights nor similar rights of any third parties by this document.
The instructions in this document must be strictly and explicitly followed by qualified and properly trained
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INCLUDING TO USERS OR OTHERS, AND DAMAGE TO OTHER PROPERTY.
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(S).
© 2014 Illumina, Inc. All rights reserved.
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To use the cBot to load 2 libraries onto a rapid flow cell, you need 1 HiSeq Rapid Duo
cBot Sample Loading Kit. Each kit contains sufficient reagents for loading 1 flow cell.
Clustering on the cBot allows 2 libraries, 1 in each lane, on the 2-lane rapid flow cell.
After template hybridization and first extension on the cBot, the remainder of the
clustering process is performed on the HiSeq 2500 or HiSeq 1500.
Kit Name
HiSeq Rapid Duo cBot Sample Loading Kit
HiSeq Rapid Duo cBot Sample Loading Kit Reagent Prep Guide
Catalog #
CT-403-2001
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Reagent Kit Overview
Reagent Kit Overview
Kit Contents and Storage Requirements
When you receive your kit, promptly store the kit components at the indicated
temperature to ensure proper performance.
Rapid Duo Kit Reagents
Component
cBot Reagent Plate
Storage
-25°C to -15°C
cBot Reagent Plate
The cBot reagent plate contains 3 rows of foil-sealed 8-tube strips filled with template
hybridization and first extension reagents. Rows 4–12 of the reagent plate are empty.
Figure 1 cBot Reagent Plate
A
B
Row 1
Row 3
Each 8-tube strip is labeled with the reagent name followed by a number. The number
indicates the row that it occupies on the reagent plate. If an 8-tube strip becomes
dislodged from the reagent plate holder, use the row number on the label to return the
tube strip to the appropriate position on the reagent plate.
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Part # 15059382 Rev. A
Kit Contents and Storage Requirements
Table 1 cBot Reagent Plate Contents
Row
Reagent
Description
HT1
Hybridization Buffer
1
AMS
Fast Amplification Mix
2
USB
Universal Sequencing Buffer
3
Empty
Empty
4
Empty
Empty
5
Empty
Empty
6
Empty
Empty
7
Empty
Empty
8
Empty
Empty
9
Empty
Empty
10
Empty
Empty
11
Empty
Empty
12
HiSeq Rapid Duo cBot Sample Loading Kit Reagent Prep Guide
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Prepare Reagent Plate
The reagent plate takes about 30 minutes to thaw using a room temperature water bath.
Alternatively, you can thaw reagents at 2°C to 8°C overnight, or up to 16 hours. After
thawing, carefully follow the steps to prepare reagents for use on the cBot.
Best Practices
} Wear a fresh pair of gloves when preparing clustering reagents.
} Hold the 96-well cBot reagent plate by the plate base to avoid dislodging reagent
tubes.
} Make sure that the tubes are securely seated in the reagent plate before and after
vortexing or inverting reagents. Loose reagent tubes can damage the cBot manifold.
NOTE
The clear plastic lid on the reagent plate protects the foil seals from being damaged or
punctured during thawing. Remove the protective lid only when necessary, such as when
you are checking that tubes are securely seated.
} After completing the reagent preparation steps, proceed to setting up the cBot for
clustering and load reagents when prompted.
Thaw the cBot Reagent Plate
1
Remove the cBot reagent plate from -25°C to -15°C storage.
2
Make sure that the tube strips are securely seated in the plate. Do not to puncture the
foil seals.
3
Place the reagent plate in a water bath containing enough room temperature
deionized water to submerge only the reagent plate base. Allow reagents to thaw for
60 minutes.
Prepare the cBot Reagent Plate
1
With a hand on top of the tubes, invert the reagent plate 5 times to mix the thawed
reagents.
2
With a hand on top of the tubes, vortex the plate for approximately 10 seconds to
dislodge trapped air bubbles.
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Part # 15059382 Rev. A
Tap the reagent plate on a hard surface approximately 10 times to collect reagent
droplets at the bottom of the tubes. Alternatively, pulse centrifuge the reagent plate.
4
Visually inspect the reagent plate to make sure that the tubes are free of air bubbles.
5
Visually inspect the tubes to make sure that the tubes are securely seated and in the
correct order as numbered on the label.
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Promptly proceed to setting up the cBot.
HiSeq Rapid Duo cBot Sample Loading Kit Reagent Prep Guide
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Prepare Reagent Plate
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Next Steps
After you have prepared the reagent plate, you are ready to prepare your libraries and
load the reagent plate when prompted by cBot software. For clustering workflow
instructions, see the cBot User Guide, part # 15006165.
Make sure that you use the compatible recipe version and cBot software version.
Library Loading Kit Version
Recipe Version
Software Version
HiSeq Rapid Duo cBot Sample
Loading Kit
Version R recipes
cBot software v1.5, or
later
After you have completed clustering on the cBot, complete the clustering process and
sequence your hybridized flow cell by performing a run in Rapid Run mode on the
HiSeq.
For workflow instructions, see the user guide for your sequencing instrument:
} HiSeq 2500 System User Guide, part # 15035786
} HiSeq 1500 System User Guide, part # 15035788
Make sure that you use the compatible version of SBS kit, cluster kit, and HiSeq Control
Software (HCS).
Library Loading Kit Version
HiSeq Rapid Duo cBot Sample
Loading Kit
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SBS Kit
Version
HiSeq Rapid
SBS Kit v2
Cluster Kit
Version
HiSeq Rapid
PE Cluster Kit v2
HiSeq Rapid SR
Cluster Kit v2
Control
Software
Version
HCS v2.2 / RTA
v1.18, or later
Part # 15059382 Rev. A
For technical assistance, contact Illumina Technical Support.
Table 2 Illumina General Contact Information
Address
5200 Illumina Way
San Diego, CA 92122 USA
Website
www.illumina.com
Email
techsupport@illumina.com
Table 3 Illumina Customer Support Telephone Numbers
Region
Contact Number
Region
North America
1.800.809.4566
Italy
Austria
0800.296575
Netherlands
Belgium
0800.81102
Norway
Denmark
80882346
Spain
Finland
0800.918363
Sweden
France
0800.911850
Switzerland
Germany
0800.180.8994
United Kingdom
Ireland
1.800.812949
Other countries
Safety Data Sheets
Safety data sheets (SDSs) are available on the Illumina website at
support.illumina.com/sds.html.
HiSeq Rapid Duo cBot Sample Loading Kit Reagent Prep Guide
Contact Number
800.874909
0800.0223859
800.16836
900.812168
020790181
0800.563118
0800.917.0041
+44.1799.534000
Technical Assistance
Technical Assistance
*15059382*
Part # 15059382 Rev. A
Illumina
San Diego, California 92122 U.S.A.
+1.800.809.ILMN (4566)
+1.858.202.4566 (outside North America)
techsupport@illumina.com
www.illumina.com