ABI PRISM 7000 Sequence Detection Systems Relative ...

More documents

Recommendations

Info

Prim 5′ 3′ Syn Are you familiar with the 7000 instrument and relative quantification No Chapter 1 Read the introduction for an overview Yes Have you designed your experiment Yes Chapter 2 Design an RQ experiment. No Have you converted total RNA to cDNA on mRNA DNA strand 5′ cDNA Reverse Primer Oligo d(T) or random hexamer 5′ cDNA 3 No Chapter 3 Reverse transcribe total RNA to cDNA. Yes Have you collected PCR data in an RQ plate document Have you analyzed data in an RQ study Yes No No Chapter 4 Chapter 5 PCR amplify the cDNA and collect data in RQ Plate documents. Analyze one or more RQ Plates in an RQ study. DRAFT September 26, 2003 3:14 pm, C1_Intro.fm Chapter 1 Introduction
Introduction 1 About the 7000 SDS Instrument Description The ABI PRISM ® 7000 Sequence Detection System (7000 SDS instrument) is a secondgeneration sequence detection instrument capable of quantitative and qualitative detection with fluorescent-based PCR chemistries. The instrument is capable of quantitative detection using real-time analysis, and qualitative detection using end-point and dissociation-curve analysis. The 7000 SDS instrument combines thermal cycling, fluorescence detection, and application-specific software. It detects accumulated polymerase chain reaction (PCR) product cycle-by-cycle, thus making quantification available immediately after completion of PCR, without the need for further process analysis. Supported Assay Types The 7000 SDS instrument allows you to perform the following assays with plates or tubes in the 96-well format: • Relative Quantification (RQ) – Determines the quantity of a single nucleic acid target sequence within an unknown sample, relative to the same sequence within a calibrator sample • Absolute Quantification (AQ) – Determines the absolute quantity of a single nucleic acid target sequence within a sample • Allelic Discrimination (AD) – Indicates the genotype of samples. • Plus/Minus – Indicates the presence or absence of a specific target sequence in a sample DRAFT September 26, 2003 3:14 pm, C1_Intro.fm For more information about the assay types, refer to the Sequence Detection Systems Chemistry Guide and the Online Help for the 7000 SDS instrument. About Relative Quantification Real-time PCR Assays Real-time PCR is the ability to monitor the progress of the PCR as it occurs. Data is collected throughout the PCR process rather than at the end of the PCR process (end-point PCR). In real-time PCR, reactions are characterized by the point in time during cycling when amplification of a target is first detected rather than the amount of target accumulated at the end of PCR. There are two types of quantitative real-time PCR: absolute and relative. Notes Relative Quantification Getting Started Guide for 7000 v1.1 1
Page 1 and 2: 5′ cDNA Reverse Primer Oligo d(T)
Page 3 and 4: Primer Extended on mRNA 5′ 3′ 5
Page 5 and 6: Chapter 5 Performing an RQ Study 33
Page 7 and 8: Preface How to Use This Guide Purpo
Page 9 and 10: Safety and EMC Compliance Informati
Page 11 and 12: Symbols on Instruments Electrical S
Page 13 and 14: Chemical Safety Chemical Hazard War
Page 15 and 16: Physical Hazard Safety Ultraviolet
Page 17: Safety and Electromagnetic Compatib
Page 21 and 22: About the Sample RQ Experiment Desc
Page 23 and 24: Materials and Equipment Description
Page 27 and 28: Specifying the Components of an RQ
Page 29 and 30: Selecting the Sequence Detection Ch
Page 31 and 32: Choosing the Probes and Primers Abo
Page 35 and 36: 5′ 3′ 5′ cDNA Reverse Primer
Page 39 and 40: Preparing the PCR Master Mix Preven
Page 41 and 42: Creating an RQ Plate Document Detec
Page 43 and 44: Specifying Thermal Cycling Conditio
Page 45 and 46: Analyzing and Viewing RQ Plate Data
Page 47 and 48: Analyzing and Viewing RQ Plate Data
Page 49 and 50: Exporting RQ Plate Documents Reanal
Page 53 and 54: Creating an RQ Study Document 5. Co
Page 55 and 56: Analyzing and Viewing the Results o
Page 57 and 58: Analyzing and Viewing the Results o
Page 59 and 60: Omitting Samples from a Study Omitt
Page 61 and 62: Exporting RQ Study Results Exportin
Page 63 and 64: References Kwok, S. and Higuchi, R.
Page 65 and 66: Creating Detectors A Before you can
Page 67 and 68: Upgrading the 7000 Software with RQ
Page 69 and 70:
Index SAMPLE DOCUMENT September 26,
Page 71 and 72:
Index SAMPLE DOCUMENT September 26,
Page 73 and 74:
DRAFT September 26, 2003 3:17 pm, G
show all

ABI PRISM 7000 Sequence Detection Systems Relative ...

Create successful ePaper yourself

Delete template?

Save as template?