Applied Biosystems 7900HT Fast Real-Time PCR System and SDS ...

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Chapter 4Operating the Instrument4. Do one of the following:• If performing a real-time run, click .• If performing an end-point run, click .The instrument tray rotates to the IN position and the instrument performs therun or plate-read.Note: Before starting the run, the instrument may pause (up to 15 minutes) toheat the heated cover to the appropriate temperature.Note: For more information on the elements of the Real-Time and Plate-Readtabs, click and see the Sequence Detection Systems Software Online Help.The following options are available during and after the completion of the run:To…See PageMonitor the progress of the run 4-27Stop the runIMPORTANT! If you must stop a run in-progress for any reason,carefully read the instructions on page 4-28 before halting the run.4-28Open the instrument tray (after the run) 4-29Analyze the run data after the run is complete 4-294-26 Applied Biosystems 7900HT Fast Real-Time PCR System and SDS Enterprise Database User GuideDRAFTSeptember 1, 2004 11:39 am, CH_Basic.fm
Running a Single Plate (Using the SDS Software)MonitoringInstrumentProgressThe SDS software displays instrument status and run progress in the Real-Time tab(real-time runs) or the Plate Read tab (end-point runs) of the respective platedocument. Figure 4-10 shows examples of the tabs during operation of the 7900HTinstrument.Real-Time Tab(Real-Time Runs)Plate Read Tab(End-Point Runs)Figure 4-10Real-Time and Plate Read Tabs of the SDS Software• Status – Displays the condition of the 7900HT instrument• Time Remaining – Displays the calculated time remaining in the runTemperature group box (Real-Time Plate Documents Only)• Block – Displays the actual temperature of the sample block module• Cover – Displays the actual temperature of the heated cover• Sample – Displays the calculated temperature of the samplesCycle group box (Real-Time Plate Documents Only)• Rep – Displays the current cycle repetition• Stage – Displays the current stage of the thermal cycling• State – Displays the current condition of the cycle stage• Step – Displays the current step being run• Time – Displays the calculated time remaining in the current stepData Collection Stamp group box (End-Point Plate Documents Only)• Pre – Displays the date that the instrument performed the pre-read.• Post – Displays the date that the instrument performed the post-read.Note: A ‘pre’ read is a plate read performed before a plate has undergone thermalcycling.Note: A ‘post’ read is a plate read performed after a plate has undergone thermalcycling.Note: For more information on the elements of the Real-Time and Plate-Read tabs,click and see the Sequence Detection Systems Software Online Help.Applied Biosystems 7900HT Fast Real-Time PCR System and SDS Enterprise Database User Guide 4-27DRAFTSeptember 1, 2004 11:39 am, CH_Basic.fm
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Applied Biosystems7900HT Fast Real-
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ContentsPrefaceHow to Use This Guid
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Chapter 4Operating the InstrumentNo
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Chapter 7Maintaining the Instrument
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Appendix D Theory of OperationFluor
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PrefaceHow to Use This GuidePurpose
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Safety and EMC Compliance Informati
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Symbols on InstrumentsSymbols on In
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General Instrument SafetyGeneral In
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Chemical Waste SafetyChemical Safet
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Physical Hazard SafetyOvervoltageRa
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Workstation SafetyWorkstation Safet
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Product Overview 11In This Chapter
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7900HT FAST Real-Time PCR SystemSec
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7900HT InstrumentInterchangeableThe
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7900HT FAST Real-Time PCR SystemBar
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Compatible ConsumablesCompatible Co
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Instrument ConnectionsFixed-Positio
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Section 1.2 Getting to Know the Sof
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SDS Software Related Files and Form
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7900HT FAST Real-Time PCR SystemMan
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Esc F1 F2 F3 F4 F5 F6 F7 F8 F9 F10
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Section 1.3 SDS Enterprise Database
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* + Enter* +Enter3Down UpNumLockSys
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About the SDS Enterprise Database S
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Esc F1 F2 F3 F4 F5 F6 F7 F8 F9 F10
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Getting Started 22In This Chapter G
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Getting StartedUsing the SDSSoftwar
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GR23967900HTPowering On the 7900HT
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Using the SDS SoftwareUsing the SDS
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Using the SDS SoftwareAbout theMess
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Basic Software Skills TutorialNotes
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Basic Software Skills TutorialExerc
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Basic Software Skills TutorialExpor
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Basic Software Skills TutorialLesso
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Basic Software Skills Tutorial3. Se
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Using SDS Plate DocumentsUsing SDS
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Using SDS Plate DocumentsWell Inspe
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Preparing a Run 33In This Chapter N
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Notes for Database UsersDescription
Page 89 and 90: Workflow OverviewWorkflow OverviewE
Page 91 and 92: Workflow OverviewAllelicDiscriminat
Page 93 and 94: Step 1 - Creating a Plate DocumentS
Page 95 and 96: Step 2 - Applying Detectors and Mar
Page 101 and 102: Step 3 - Configuring the Plate Docu
Page 103 and 104: Step 5 - Programming the Plate Docu
Page 109 and 110: Step 6 - Saving the Plate Document
Page 111 and 112: Step 7 - Creating a Plate Document
Page 113 and 114: Step 9 - Running the Plate on the 7
Page 115 and 116: Operating the Instrument 44In This
Page 117 and 118: Notes for Database UsersDescription
Page 119 and 120: Section 4.1 Consumable PreparationS
Page 121 and 122: Compatible ConsumablesTable 4-1Comp
Page 123 and 124: Preparing Optical Plates for Usea.
Page 125 and 126: Preparing TaqMan Low Density Arrays
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Page 139: Running a Single Plate (Using the S
Page 143 and 144: After the RunAfter the RunUser Acce
Page 145 and 146: Section 4.3 Automated OperationSect
Page 147 and 148: Operating the Software with an SDS
Page 149 and 150: Operating the Software without an S
Page 155 and 156: Running Plates Using the Automation
Page 157 and 158: Running Plates Using the Automation
Page 159 and 160: Analyzing End-Point Data 55In This
Page 161 and 162: Notes for Database UsersNotes for D
Page 163 and 164: Section 5.1 Allelic DiscriminationS
Page 165 and 166: OverviewTable 5-1Signal and Sequenc
Page 167 and 168: Before You BeginBefore You BeginUsi
Page 169 and 170: Opening the Run DataOpening the Run
Page 171 and 172: Calling and Scrutinizing Allelic Di
Page 177 and 178: After the AnalysisSaving theResults
Page 179 and 180: Analyzing Real-Time Data 66In This
Page 181 and 182: Notes for Database UsersDescription
Page 183 and 184: Section 6.1 Absolute Quantification
Page 185 and 186: OverviewAlgorithmicManipulation ofR
Page 187 and 188: Analysis ChecklistAnalysis Checklis
Page 189 and 190: Analyzing the DataAnalyzing the Dat
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Viewing ResultsViewing ResultsViewi
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Section 6.2 Relative Quantification
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OverviewAlgorithmic Manipulation of
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OverviewAutomatic Outlier RemovalAb
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Getting StartedGetting StartedUsing
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Options for Analyzing Relative Quan
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Creating the Studyd. In the Plate Q
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Analyzing the Study• Manual Ct -
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Analyzing the StudySpecifying the A
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Viewing ResultsViewing ResultsDispl
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Viewing ResultsAbout DownArrowsDown
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After the AnalysisAfter the Analysi
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Section 6.3 Dissociation Curve Anal
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Before You BeginExample ResultsFigu
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Opening the Run DataOpening the Run
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Determining T m Values for the Anal
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Section 6.4 Procedure ReferenceSect
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Setting the Baseline and Threshold
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Eliminating Outlying AmplificationE
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Eliminating Outlying Amplification6
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After the AnalysisSaving theResults
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Maintaining the Instrument 77In Thi
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Notes for Database UsersDescription
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Section 7.1 Maintaining the 7900HT
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Replacing the Sample BlockMaterials
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Replacing the Sample Block9. Loosen
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7900HT FAST Real-Time PCR SystemRep
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Changing the Plate Adapter4. Place
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Decontaminating the Sample BlockCle
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Performing a Background RunPreparin
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Performing a Background RunAnalyzin
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Performing a Pure Dye RunAfter a ru
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Performing a Pure Dye Run• Fast 9
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Performing a Pure Dye RunAnalyzing
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Adding Custom Dyes to the Pure Dye
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Adding Custom Dyes to the Pure Dye
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Verifying Instrument PerformanceTab
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Verifying Instrument PerformancePre
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Section 7.2 Maintaining the Plate H
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Adjusting the Sensitivity of the Pl
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Adjusting the Sensitivity of the Pl
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7900HT FAST Real-Time PCR SystemAli
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Aligning the Plate Handler5. Using
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Aligning the Plate Handler7. Using
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Aligning the Plate Handler13. Click
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7900HT FAST Real-Time PCR SystemAli
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Aligning the Fixed-Position Bar Cod
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Section 7.3 Maintaining the Compute
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Maintaining the SDS softwareSeveral
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Troubleshooting 88In This Chapter T
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Troubleshooting TableTable 8-1Troub
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Low Precision or IrreproducibilityL
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Low Precision or IrreproducibilityD
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Background RunsBackground RunsBackg
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Pure Dye Runs6. Repeat step 4 until
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End-Point Runs (Allelic Discriminat
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Software and 7900HT InstrumentTable
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Zymark Twister Microplate Handler a
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TaqMan Low Density ArrayTable 8-6Tr
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Software ReferenceAAIn This Appendi
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Importing Plate Document Setup Tabl
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Setup Table Files1. File VersionDes
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Setup Table Files6. Detectors ListD
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Setup Table Files9. Markers List (A
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Setup Table FilesFormat(for a singl
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Setup Table FilesFormat(for a singl
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Setup Table FilesExample Code A-16A
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Exporting Plate Document Data4. In
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Operating the SDS Software from a C
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Using the Search ToolLogical Operat
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Connecting SDS Software to the Data
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Designing TaqMan Reagent-BasedAssay
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Assay Development Guidelines• For
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Design Tips for Allelic Discriminat
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Kits, Reagents, and ConsumablesCCIn
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Consumables and DisposablesConsumab
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Instrument Maintenance and Verifica
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Custom Oligonucleotide SynthesisTab
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Theory of OperationDDIn This Append
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Fluorescent-Based ChemistriesBasics
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Real-Time Data AnalysisPhase 2: Lin
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Real-Time Data AnalysisSignificance
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A similar equation for the endogeno
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Limited Warranty StatementEEWarrant
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BibliographyGeneral Paperson TaqMan
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Klein, D., Janda, P., Steinborn, R.
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Dolken, L., F. , Schuler, and G. ,
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AllelicDiscriminationVirtanen M., H
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Glossary.sdd.sdm.sds.sdt∆C T∆
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ackgroundbaselinecalibratorcDNA(com
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minor groovebinder (MGB)multicompon
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R nrunsessionsingle nucleotidepolym
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IndexSymbols- 6-31↑ 6-33Numerics7
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fluorescent, common sources 8-7isol
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adjusting step parameters 3-21confi
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sample block locking bar 7-8sample
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Worldwide Sales and SupportApplied
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