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illumina MiniSeq System Manual

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MiniSeq
System Guide
Document # 1000000002695 v02 Material # 20014309
March 2018
For Research Use Only. Not for use in diagnostic procedures.
ILLUMINA PROPRIETARY

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  • Page 1 MiniSeq System Guide Document # 1000000002695 v02 Material # 20014309 ILLUMINA PROPRIETARY March 2018 For Research Use Only. Not for use in diagnostic procedures.
  • Page 2 MiniSeq System Guide 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.
  • Page 3 Document Date Description of Change Material # 20014309 March Added information about the Illumina Proactive monitoring service in Document # 2018 the Configure Analysis Settings section. 1000000002695 v02 Removed the default user name and password required to log on to the operating system.

  • Page 4: Table Of Contents

    Table of Contents Chapter 1 Overview Introduction Additional Resources Instrument Components Sequencing Consumables Overview Preinstalled Databases and Genomes Chapter 2 Getting Started Starting the Instrument Customize System Settings Configure Analysis Settings User-Supplied Consumables and Equipment Chapter 3 Sequencing Introduction Sequencing Workflow Prepare Consumables Prepare Libraries for Sequencing Set Up a Sequencing Run...
  • Page 5 MiniSeq System Guide Appendix C Output Files Sequencing Output Files Sequencing Output Folder Structure Analysis Input File Requirements Index Technical Assistance Document # 1000000002695 v02 Material # 20014309 For Research Use Only. Not for use in diagnostic procedures.

  • Page 6: Chapter 1 Overview

    High-quality sequencing—The MiniSeq System enables small genome, amplicon, targeted enrichment, and RNA sequencing using low library volumes. MiniSeq system software—The MiniSeq System includes a suite of integrated software that controls instrument operations, processes images, and generates base calls. The suite includes on-instrument data analysis software and data transfer tools for analysis using other options, such as BaseSpace.

  • Page 7: Instrument Components

    # 1000000002702) Instrument Components The MiniSeq System includes a touch screen monitor, a status bar, a flow cell compartment, and a reagent compartment. Figure 1 Instrument Components A Touch screen monitor—Enables on-instrument configuration and setup using the control software interface.
  • Page 8 Do not place objects on the instrument near the flow cell compartment. Reagent Compartment Setting up a sequencing run on the MiniSeq System requires access to the reagent compartment to load run consumables and empty the spent reagents bottle. Figure 3 Reagent Compartment A Spent reagents bottle—Includes a threaded cap to prevent spills when carrying.
  • Page 9 The instrument software suite includes integrated applications that perform sequencing runs and on- instrument analysis. MiniSeq Control Software—The control software guides you through the steps to set up a sequencing run, controls instrument operations, and shows an overview of run statistics as the run progresses.

  • Page 10: Sequencing Consumables Overview

    Select Acknowledge to accept the message and Close to close the dialog box. Sequencing Consumables Overview Performing a sequencing run on the MiniSeq System requires a single-use MiniSeq Kit. Each kit includes a flow cell and the reagents required for a sequencing run.
  • Page 11 Handle used reagents as chemical waste and discard in accordance with applicable regional, national, and local laws and regulations. For additional environmental, health, and safety information, see the SDS at support.illumina.com/sds.html. Store the reagent cartridge at -25°C to -15°C until use. For more information, see...

  • Page 12: Preinstalled Databases And Genomes

    MiniSeq System Guide Position Description 13, 14, and 15 Reserved for optional custom primers Load libraries Removable Reservoir in Position #9 The prefilled reagent cartridge includes a denaturation reagent in position #9 that contains formamide. To facilitate safe disposal of any unused reagent after the sequencing run, this reservoir is removable. For more...

  • Page 13: Chapter 2 Getting Started

    Figure 8 Power Button Located on Front of Instrument Wait until the operating system has finished loading. The MiniSeq Control Software launches and initializes the system automatically. The Home screen opens after the initialization step is complete. Wait for the system to load, and then log on to the operating system. If necessary, consult your facility administrator for the user name and password.
  • Page 14 MiniSeq System Guide Purging consumables at end of run. Skipping pre-run check confirmation to start the run automatically. Using the on-screen keyboard for run setup steps. Enabling audio indicators. Performing a simulated run. Customize Instrument Identification From the Manage Instrument screen, select System Customization.

  • Page 15: Configure Analysis Settings

    When this field is blank, exiting Demo Mode does not require a PIN. Select OK to restart the software in Demo Mode. After restart, the MiniSeq Control Software banner reads Demonstration Only. Exit Demo Mode From the Manage Instrument screen, select System Customization.
  • Page 16 The name of the setting in the software interface might be different from the name in this guide, depending on the version of MiniSeq Control Software in use. With this setting turned on, instrument performance data are sent to Illumina. This data helps Illumina troubleshoot more easily and detect potential failures, enabling proactive maintenance and maximizing instrument uptime.
  • Page 17 MiniSeq System Guide With this setting turned on, instrument performance data are sent to Illumina. This data helps Illumina troubleshoot more easily and detect potential failures, enabling proactive maintenance and maximizing instrument uptime. For more information on the benefits of this service, see Illumina Proactive Technical Note (document # 1000000052503) .

  • Page 18: User-Supplied Consumables And Equipment

    The name of the setting in the software interface might be different from the name in this guide, depending on the version of MiniSeq Control Software in use. With this setting turned on, instrument performance data are sent to Illumina. This data helps Illumina troubleshoot more easily and detect potential failures, enabling proactive maintenance and maximizing instrument uptime.
  • Page 19 MiniSeq System Guide User-Supplied Consumables for Instrument Maintenance Consumable Supplier Purpose NaOCl, 5% Sigma-Aldrich, catalog # 239305 Performing a manual post-run wash; (sodium hypochlorite) (or laboratory-grade equivalent) diluted to 0.12% Tween 20 Sigma-Aldrich, catalog # P7949 Performing a manual instrument wash;...
  • Page 20 MiniSeq System Guide Document # 1000000002695 v02 Material # 20014309 For Research Use Only. Not for use in diagnostic procedures.

  • Page 21: Chapter 3 Sequencing

    Monitor Run Progress Automatic Post-Run Wash Remove Used Reservoir from Position #9 Introduction To perform a sequencing run on the MiniSeq System, prepare run consumables and then follow the software prompts to set up the sequencing run. Workflow Overview Cluster Generation During cluster generation, single DNA molecules are bound to the surface of the flow cell, and then amplified to form clusters.

  • Page 22: Sequencing Workflow

    BaseSpace or BaseSpace Onsite, or Local Run Manager—Before setting up the run on the instrument, assign a run name and specify run parameters using the BaseSpace Prep tab or the Local Run Manager software. After preparing consumables, select the run name from the MiniSeq Control Software, and then load consumables for the run.
  • Page 23 Prepare a new reagent cartridge: thaw and inspect. Prepare a new flow cell: bring to room temperature, unwrap, and inspect. Denature and dilute libraries. See MiniSeq System Denature and Dilute Libraries Guide (document # 1000000002697) . Load the library dilution onto the reagent cartridge in reservoir #16.

  • Page 24: Prepare Consumables

    MiniSeq System Guide Prepare Consumables Prepare the Reagent Cartridge Remove the reagent cartridge from -25°C to -15°C storage. Thaw reagents using the following water bath options. Do not submerge the cartridge. When thawed, dry the base before proceeding. Method Time to Thaw Stability Limit 37°C water bath...

  • Page 25: Prepare Libraries For Sequencing

    For more information, see MiniSeq System Denature and Dilute Libraries Guide (document # 1000000002697) . The loading volume for the MiniSeq System is 500 µl at a loading concentration of 1.8 pM. However, loading concentration can vary depending on library preparation and quantification methods.
  • Page 26 MiniSeq System Guide Load the Flow Cell Open the flow cell compartment door. Press the release button to the right of the flow cell latch. Figure 11 Open Flow Cell Latch If present, remove the used flow cell from a previous run.
  • Page 27 MiniSeq System Guide If present, remove the used reagent cartridge. NOTE To facilitate safe disposal of unused reagent containing formamide, the reservoir in position #9 is Remove Used Reservoir from Position #9 on page removable. See Slide the reagent cartridge into the reagent compartment until the cartridge stops.
  • Page 28 MiniSeq System Guide Figure 16 Seal the Spent Reagents Bottle Discard the contents in accordance with applicable standards. WARNING This set of reagents contains potentially hazardous chemicals. Personal injury can occur through inhalation, ingestion, skin contact, and eye contact. Wear protective equipment, including eye protection, gloves, and laboratory coat appropriate for risk of exposure.
  • Page 29 MiniSeq System Guide [Optional] If you are using custom primers, select the checkbox for the primers used. Read 1—Custom primer for Read 1. Index 1—Custom primer for Index 1. Index 2—Custom primer for Index 2. Read 2—Custom primer for Read 2.
  • Page 30 MiniSeq System Guide Load the Flow Cell Open the flow cell compartment door. Press the release button to the right of the flow cell latch. Figure 17 Open Flow Cell Latch If present, remove the used flow cell from a previous run.
  • Page 31 MiniSeq System Guide If present, remove the used reagent cartridge. NOTE To facilitate safe disposal of unused reagent containing formamide, the reservoir in position #9 is Remove Used Reservoir from Position #9 on page removable. See Slide the reagent cartridge into the reagent compartment until the cartridge stops.
  • Page 32 MiniSeq System Guide Figure 22 Seal the Spent Reagents Bottle Discard the contents in accordance with applicable standards. WARNING This set of reagents contains potentially hazardous chemicals. Personal injury can occur through inhalation, ingestion, skin contact, and eye contact. Wear protective equipment, including eye protection, gloves, and laboratory coat appropriate for risk of exposure.

  • Page 33: Monitor Run Progress

    MiniSeq System Guide NOTE Do not open the reagent compartment door or the flow cell compartment door during the automated check or during the sequencing run. Review Automated Check Review the automated check results. To stop a check in progress, select Cancel.
  • Page 34 BaseSpace and an additional output folder location, a BaseSpace icon and a standalone icon appear. Run Copy Service The MiniSeq System Software Suite includes a Run Copy Service. As RTA generates files, the service copies files to the specified output folder location.

  • Page 35: Automatic Post-Run Wash

    MiniSeq System Guide If data transfer is interrupted during the run, data are stored temporarily on the instrument computer. When the connection is restored, data transfer resumes automatically during the run. If the connection is not restored before the run ends, manually move data to the preferred location.
  • Page 36 Handle used reagents as chemical waste and discard in accordance with applicable regional, national, and local laws and regulations. For additional environmental, health, and safety information, see the SDS at support.illumina.com/sds.html. Document # 1000000002695 v02 Material # 20014309...

  • Page 37: Chapter 4 Maintenance

    Software Updates on page Preventive Maintenance Illumina recommends that you schedule a preventive maintenance service each year. If you are not under a service contract, contact your Territory Account Manager or Illumina Technical Support to arrange for a billable preventive maintenance service.
  • Page 38 MiniSeq System Guide Figure 26 Wash Cartridge Prepare for a Manual Post-Run Wash Combine the following volumes to result in 0.12% NaOCl. 5% NaOCl (31 µl) Laboratory-grade water (1269 µl) Add 1.3 ml of 0.12% NaOCl to the wash cartridge. The correct reservoir is equivalent to position #31 on the prefilled reagent cartridge.
  • Page 39 MiniSeq System Guide Prepare for a Quick Wash Combine the following volumes to result in a 0.05% Tween 20 wash solution. 100% Tween 20 (20 µl) Laboratory-grade water (40 ml) Add 40 ml wash solution to the wash cartridge. The correct reservoir is equivalent to position #40 on the prefilled reagent cartridge.

  • Page 40: Software Updates

    MiniSeq Service Software Sequencing Analysis Viewer BaseSpace Broker Software release notes are available on the MiniSeq System support page on the Illumina website. You can install software updates automatically using an internet connection or manually from a network or USB location.
  • Page 41 If a firmware update is included, an automatic restart of the system is required after the firmware is updated. Manual Software Update Download the System Suite installer from the Illumina website and save it to a network location. Alternatively, copy the software installation file to a portable USB drive.
  • Page 42 MiniSeq System Guide Document # 1000000002695 v02 Material # 20014309 For Research Use Only. Not for use in diagnostic procedures.

  • Page 43: Appendix A Troubleshooting

    Lane log files are updated throughout the run. (*LaneLog*.txt) Troubleshooting Resources For technical questions, visit the MiniSeq System support pages on the Illumina website. The support pages provide access to documentation, downloads, and frequently asked questions. Log in to your MyIllumina account for access to support bulletins.

  • Page 44: Automatic Check Errors

    MiniSeq System Guide Process Status The MiniSeq Control Software lists the status of at least 3 runs in the system Temp folder. From the Manage Instrument screen, select Process status. For each run name, the system lists the status of the following components: Real-Time Analysis (RTA)—Based on the processing of BCL files...

  • Page 45: Rta Errors

    RTA Errors To troubleshoot RTA errors, first check the RTA error log, which is stored in the RTALogs folder. This file is not present for successful runs. Include the error log when reporting issues to Illumina Technical Support. Error Handling RTA2 creates log files and writes them to the RTALogs folder.

  • Page 46: Rehybridization Workflow

    Required Consumables A rehybridization run requires a new MiniSeq reagent cartridge regardless of when the run was stopped. End the Current Run Select End Run. When prompted to confirm the command, select Yes.

  • Page 47: System Check

    Select Next to proceed to the pre-run check and start the run. System Check A system check is not required for normal operation or instrument maintenance. However, an Illumina Technical Support representative might ask you to perform a system check for troubleshooting purposes.
  • Page 48 MiniSeq System Guide Starting a system check automatically closes the control software and launches the MiniSeq Service Software. The service software launches and opens to the Load screen, which is configured to use the advanced loading option. Figure 29 Available System Checks After loading consumables, the Select screen opens, which lists available system checks.
  • Page 49 MiniSeq System Guide If a used flow cell is not already on the instrument, load a used flow cell. b Empty the spent reagents bottle and return it to the instrument. Load the wash cartridge. Select Load. The software moves the flow cell and wash cartridge into position.

  • Page 50: Network Configuration Settings

    MiniSeq System Guide Network Configuration Settings Network settings are configured during installation. If the system has to be reconfigured, you can change settings or reset settings on the Network Configuration screen. Configuration settings include the IP address, domain name server (DNS) address, and domain name.

  • Page 51: Custom Genomes

    CAUTION Do not relocate the instrument. Moving the instrument improperly can affect the optical alignment and compromise data integrity. If you have to relocate the instrument, contact your Illumina representative. Document # 1000000002695 v02 Material # 20014309 For Research Use Only. Not for use in diagnostic procedures.
  • Page 52 MiniSeq System Guide Document # 1000000002695 v02 Material # 20014309 For Research Use Only. Not for use in diagnostic procedures.

  • Page 53: Appendix B Real-Time Analysis

    The MiniSeq System uses an implementation of Real-Time Analysis called RTA2. The system control software and RTA2 communicate through a web HTTP interface and shared memory files. If RTA2 is terminated, processing does not resume and run data are not saved.

  • Page 54: Real-Time Analysis Workflow

    MiniSeq System Guide Real-Time Analysis Workflow Template generation Maps cluster locations. Registration and intensity Records the location of each cluster on the flow cell and determines an intensity value for extraction each cluster. Phasing correction Corrects the effects of phasing and prephasing.
  • Page 55 Base Calling Base calling determines a base (A, C, G, or T) for every cluster of a given tile at a specific cycle. The MiniSeq System uses 2-channel sequencing, which requires only 2 images to encode the data for 4 DNA bases, 1 from the red channel and 1 from the green channel.
  • Page 56 MiniSeq System Guide For 2-channel analysis, RTA2 uses a population-based system to determine the chastity of a base call. Clusters pass filter (PF) when no more than 1 base call in the first 25 cycles has an unacceptable chastity value. Clusters that do not pass filter are not base called in future cycles.

  • Page 57: Appendix C Output Files

    Appendix C  Output Files Output Files Sequencing Output Files Sequencing Output Folder Structure Analysis Input File Requirements Sequencing Output Files File Type File Description, Location, and Name Base call files Each tile analyzed is included in a base call file, aggregated in a file for each cycle. The aggregated file contains the base call and encoded quality score for every cluster.

  • Page 58: Analysis Input File Requirements

    MiniSeq System Guide L001—Focus images InstrumentAnalyticsLogs—Log files describing instrument analytics steps InterOp—Binary files used by Sequencing Analysis Viewer Logs—Log files describing operational steps Recipe—Run-specific recipe file named with reagent cartridge ID RTALogs—Log files describing analysis steps RTAComplete.xml RTAConfiguration.xml RunInfo.xml RunNotes.xml RunParameters.xml...

  • Page 59: Index

    14 errors and warnings 4 reagent cartridge 6 status 4 sequencing runs 13 Illumina Proactive monitoring service 11-13 user-supplied 13 Document # 1000000002695 v02 Material # 20014309 For Research Use Only. Not for use in diagnostic procedures.
  • Page 60 MiniSeq System Guide imaging compartment 2 purge consumables 9 imaging, 2-channel sequencing 50 indexing considerations 51 instrument configuration settings 45 Q-scores 51 power button 4 quality tables 51 start up 8 instrument maintenance consumables 14 instrument wash 32 read length 16...
  • Page 61 MiniSeq System Guide automatic update 35 pre-run check 24, 28 check for update 9 reagent cartridge 19, 22, 26 configuration settings 45 run duration 16 image analysis, base calling 4 run metrics 28 initialization 8 sequencing 49 instrument control 4...

  • Page 62: Technical Assistance

    Other countries +44.1799.534000 Safety data sheets (SDSs)—Available on the Illumina website at support.illumina.com/sds.html. Product documentation—Available for download in PDF from the Illumina website. Go to support.illumina.com, select a product, then select Documentation & Literature. Document # 1000000002695 v02 Material # 20014309...

  • Page 63: Document # 1000000002695 V02 Material # 20014309

    Document # 1000000002695 v02 Material # 20014309 Illumina 5200 Illumina Way 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 For Research Use Only. Not for use in diagnostic procedures. © 2018 Illumina, Inc. All rights reserved.

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