HiSeq Rapid SBS Kit v2 Reagent Prep Guide ®
Transcription
HiSeq Rapid SBS Kit v2 Reagent Prep Guide ®
HiSeq® Rapid SBS Kit v2 Reagent Prep Guide FOR RESEARCH USE ONLY Reagent Kit Overview Kit Contents and Storage Requirements Number of Cycles Prepare SBS Reagents Next Steps Reagent Positions Technical Assistance ILLUMINA PROPRIETARY Part # 15058772 Rev. A Catalog # FC-402-9003DOC November 2014 3 4 5 6 7 8 This document and its contents are proprietary to Illumina, Inc. and its affiliates ("Illumina"), and are intended solely for the contractual use of its customer in connection with the use of the product(s) described herein and for no other purpose. This document and its contents shall not be used or distributed for any other purpose and/or otherwise communicated, disclosed, or reproduced in any way whatsoever without the prior written 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 personnel in order to ensure the proper and safe use of the product(s) described herein. All of the contents of this document must be fully read and understood prior to using such product(s). FAILURE TO COMPLETELY READ AND EXPLICITLY FOLLOW ALL OF THE INSTRUCTIONS CONTAINED HEREIN MAY RESULT IN DAMAGE TO THE PRODUCT(S), INJURY TO PERSONS, INCLUDING TO USERS OR OTHERS, AND DAMAGE TO OTHER PROPERTY. ILLUMINA DOES NOT ASSUME ANY LIABILITY ARISING OUT OF THE IMPROPER USE OF THE PRODUCT(S) DESCRIBED HEREIN (INCLUDING PARTS THEREOF OR SOFTWARE) OR ANY USE OF SUCH PRODUCT(S) OUTSIDE THE SCOPE OF THE EXPRESS WRITTEN LICENSES OR PERMISSIONS GRANTED BY ILLUMINA IN CONNECTION WITH CUSTOMER'S ACQUISITION OF SUCH PRODUCT (S). FOR RESEARCH USE ONLY © 2014 Illumina, Inc. All rights reserved. Illumina, 24sure, BaseSpace, BeadArray, BlueFish, BlueFuse, BlueGnome, cBot, CSPro, CytoChip, DesignStudio, Epicentre, GAIIx, Genetic Energy, Genome Analyzer, GenomeStudio, GoldenGate, HiScan, HiSeq, HiSeq X, Infinium, iScan, iSelect, ForenSeq, MiSeq, MiSeqDx, MiSeq FGx, NeoPrep, Nextera, NextBio, NextSeq, Powered by Illumina, SeqMonitor, SureMDA, TruGenome, TruSeq, TruSight, Understand Your Genome, UYG, VeraCode, verifi, VeriSeq, the pumpkin orange color, and the streaming bases design are trademarks of Illumina, Inc. and/or its affiliate(s) in the U.S. and/or other countries. All other names, logos, and other trademarks are the property of their respective owners. To perform a rapid run on the HiSeq 2500 or HiSeq 1500, you need at least 1 HiSeq Rapid SBS Kit v2. Kit Name Catalog # HiSeq Rapid SBS Kit v2 (500 cycles) FC-402-4023 HiSeq Rapid SBS Kit v2 (200 cycles) FC-402-4021 HiSeq Rapid SBS Kit v2 (50 cycles) FC-402-4022 Each kit includes sequencing reagents used on the HiSeq and a set of funnel caps. The reagents are sufficient for sequencing 1 flow cell. Kit Component Quantity Rapid SBS Kit v2 (Box 1 of 2) 1 Rapid SBS Kit v2 (Box 2 of 2) 1 Set of 8 funnel caps 1 SBS Kit Features Reagent kits include the following features: } Reagents are provided in 250 ml bottles that load directly onto reagent racks. } Reagent labels are color-coded to reduce errors while loading reagents. } Each reagent contains a barcode for tracking and a human-readable label. NOTE Scan the reagent kit ID from box 1 during the run setup steps before loading reagents. HiSeq Rapid SBS Kit v2 Reagent Prep Guide 3 Reagent Kit Overview Reagent Kit Overview Kit Contents and Storage Requirements HiSeq SBS reagents are shipped in 2 boxes. One box is shipped at room temperature and the other box is shipped on dry ice. When you receive your kit, promptly store the kit components at the indicated temperature to ensure proper performance. NOTE Reagents are sensitive to light. Store reagents in a dark place. Reagents are labeled with a 3-letter acronym plus a numeric designator that indicates the kit size. For example, if you are using CRM-50, the CRM was provided in a 50-cycle kit. SBS Kit Box 1 Kit Component Quantity Storage Description CWM 1 15°C to 30°C Cleavage Wash Mix PW1 1 15°C to 30°C Wash Buffer USB 2 15°C to 30°C Universal Sequencing Buffer Storage Description SBS Kit Box 2 Kit Component Quantity IMT 1 -25°C to -15°C Incorporation Master Mix CRM 1 -25°C to -15°C Cleavage Reagent Mix USM 1 -25°C to -15°C Universal Scan Mix 4 Part # 15058772 Rev. A Rapid SBS kits contain a sufficient volume of reagents for performing dual-indexed runs: } One 500-cycle SBS kit provides reagents for up to 525 cycles of sequencing. } One 200-cycle SBS kit provides reagents for up to 225 cycles of sequencing. } One 50-cycle SBS kit provides reagents for up to 74 cycles of sequencing. } For runs greater than the maximum number of cycles in a kit and to maximize reagent use for other read lengths, use multiple 50-cycle kits. The following table lists the number of cycles performed for dual-indexed runs of various lengths. For dual-indexed paired-end runs, 7 chemistry-only cycles are calculated in the total number of cycles. Run Type Dual-Indexed Paired-End Dual-Indexed Single-Read HiSeq Rapid SBS Kit v2 Reagent Prep Guide Run Length Run Cycles Total Cycles 2 x 250 2 x 100 2 x 92 2 x 75 2 x 50 2 x 35 1 x 250 1 x 100 1 x 92 1 x 75 1 x 50 1 x 35 251+8+(7)+8+251 101+8+(7)+8+101 93+8+(7)+8+93 76+8+(7)+8+76 51+8+(7)+8+51 36+8+(7)+8+36 251+8+8 101+8+8 93+8+8 76+8+8 51+8+8 36+8+8 525 225 209 175 125 95 267 117 109 92 67 52 5 Number of Cycles Number of Cycles Prepare SBS Reagents SBS reagents are loaded onto the instrument at the beginning of the run. Before setting up the run, use the following instructions to thaw and inspect reagents. Thaw Reagents 1 Remove CRM, IMT, and USM from -25°C to -15°C storage. 2 Thaw reagents in a room temperature deionized water bath for about 90 minutes. Alternatively, thaw reagents at 2°C to 8°C for up to 16 hours. NOTE Always replace your gloves after handling CRM. 3 Invert each bottle several times to mix. 4 Inspect the reagents for ice crystals and precipitate. Make sure that the reagents have thawed completely and precipitate in the USM bottle is cleared. 5 Set CRM, IMT, and USM aside at 2°C to 8°C until you are ready to load them onto the instrument. 6 Use CWM, PW1, and USB directly from storage. 6 Part # 15058772 Rev. A After you have prepared SBS reagents, you are ready to load them onto the instrument when prompted by the control software. For reagent loading and sequencing 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 cluster kit and control software. SBS Kit Version Cluster Kit Version Control Software HiSeq Rapid SBS Kit v2 HiSeq Rapid PE Cluster Kit v2 HiSeq Rapid SR Cluster Kit v2 HiSeq Rapid Duo cBot Sample Loading Kit HCS v2.2 / RTA v1.18, or later HiSeq Rapid SBS Kit v2 Reagent Prep Guide 7 Next Steps Next Steps Reagent Positions Reagent bottles have color-coded labels that match their positions in the reagent racks. Load reagents as described in the user documentation for your sequencing instrument. To prepare for the post-run rinse at the end of a sequencing run, load 25 ml of PW1 or laboratory-grade water in positions 2 and 4. Figure 1 SBS Reagent Rack Table 1 SBS Reagent Positions Position Reagent 1 2 3 4 5 6 7 8 8 IMT PW1 USM PW1 USB USB CRM CWM Description Incorporation Mastermix 25 ml of PW1 or laboratory-grade water Universal Scan Mix 25 ml of PW1 or laboratory-grade water Universal Sequencing Buffer Universal Sequencing Buffer Cleavage Reagent Mastermix Cleavage Wash Mix Part # 15058772 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 [email protected] 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 Contact Number 800.874909 0800.0223859 800.16836 900.812168 020790181 0800.563118 0800.917.0041 +44.1799.534000 Safety Data Sheets Safety data sheets (SDSs) are available on the Illumina website at support.illumina.com/sds.html. Product Documentation Product documentation in PDF is available for download from the Illumina website. Go to support.illumina.com, select a product, then click Documentation & Literature. HiSeq Rapid SBS Kit v2 Reagent Prep Guide Technical Assistance Technical Assistance *15058772* Part # 15058772 Rev. A Illumina San Diego, California 92122 U.S.A. +1.800.809.ILMN (4566) +1.858.202.4566 (outside North America) [email protected] www.illumina.com