Mapping 250K/500K SNP assay

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OTHER EQUIPMENT REQUIRED 230 GeneChip ® Mapping 500K 96-Well Plate Protocol Table A.6 Other Equipment Required to Run the GeneChip ® Mapping 500K 96-Well Plate Protocol Equipment Quantity Manufacturer/ Distributor Cooler StrataCooler ® Lite Benchtop StrataCooler ® II Benchtop 2 Stratagene Part Number Laboratory Location 400012 Pre-PCR and Main Lab 400002 (blue) 400008 (red) Cooling chamber single gold block 2 double Diversified CHAM-1000 1 single in double gold block and 1 single Biotech CHAM-1020 Pre-PCR; enough for 4 plates in Main Lab Heat block (required only if using tubes for Stage 9: Target Hybridization) 2 Fisher Scientific 11-718-9 Main Lab Ice bucket — 2 — — Pre-PCR and Main Lab Jitterbug, 115V AC 1 Boekel Scientific 130000 Main Lab Manifold, QIAvac multiwell unit 1 QIAGEN 9014579 Main Lab Microcentrifuge, PicoFuge ® (maximum rotation 6000 rpm) Pipet-Lite, Magnetic-Assist single channel P10 Pipet-Lite, Magnetic-Assist single channell P20 Pipet-Lite, Magnetic-Assist single channel P100 Pipet-Lite, Magnetic-Assist single channel P200 Pipet-Lite, Magnetic-Assist single channel P1000 Pipette, 12-channel P20 (accurate to within ± 5%) 2 Stratagene 400550 Pre-PCR and Main Lab 2 Rainin L-10 Main Lab 2 Rainin L-20 Pre-PCR and Main Lab 2 Rainin L-100 Main Lab 2 Rainin L-200 Pre-PCR and Main Lab 2 Rainin L-1000 Main Lab 2 Rainin P/N L12-20 Pre-PCR and Main Lab
appendix A | Reagents, Equipment, and Consumables Required for 96-Well Plate Protocol 231 Table A.6 Other Equipment Required to Run the GeneChip ® Mapping 500K 96-Well Plate Protocol Equipment Quantity Manufacturer/ Distributor Pipette, 12-channel P100 2 Rainin P/N L12-100 Pre-PCR and Main Lab Pipette, 12-channel P200 2 Rainin P/N L12-200 Pre-PCR and Main Lab Plate, UV Star Transparent, 96-well 3 E&K Scientific EK-25801 Main Lab Plate Centrifuge, multipurpose 2 Eppendorf 5804 or 5810 Pre-PCR and Main Lab Plate stand (optional) 1 Diversified Biotech Spectrophotometer, high throughput microplate spectrophotometer Thermal Cyclers – see Table A.7 on page 232 Timers (required only if using tubes for Stage 9: Target Hybridization) Vacuum regulator Protocol requires 600 mb vacuum. If your lab does not have an internally regulated vacuum source, this regulator is strongly recommended. 1 Molecular Devices Part Number Laboratory Location WPST-1000 Main Lab SpectraMax Plus 384 Main Lab 4 — — Main Lab 1 QIAGEN 19530 Main Lab Vortexer, for plates and tubes 2 VWR 58816-12 Pre-PCR and Main Lab
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GeneChip ® Mapping 500K Assay Manu
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Contents CHAPTER 1 Overview 1 ABOUT
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contents v Equipment and Consumable
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contents vii Reagents Required 118
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contents ix CHAPTER 8 Troubleshooti
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contents xi STEP 4: PCR 257 Reagent
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Chapter 1 Overview
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About This Manual This manual is a
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chapter 1 | Overview 5 LD patterns
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Figure 1.1 GeneChip ® Mapping Assa
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chapter 1 | Overview 9 Gilman, B.,
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chapter 1 | Overview 11 23. Schaid,
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chapter 1 | Overview 13 35. Hu, N.,
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chapter 1 | Overview 15 50. Bignell
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chapter 1 | Overview 17 64. Gabriel
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Chapter 2 Laboratory Setup
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Introduction to Laboratory Setup Pl
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Main Lab Pre-PCR Clean Room chapter
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Chapter 3 Genomic DNA General Requi
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Introduction This chapter describes
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Sources of Human Genomic DNA chapte
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chapter 3 | Genomic DNA General Req
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Chapter 4 96-Well Plate Protocol
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ABOUT THIS PROTOCOL 96-Well Plate P
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chapter 4 | 96-Well Plate Protocol
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PREPARING THE WORK AREA FOR EACH ST
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PROGRAM YOUR THERMAL CYCLERS chapte
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EQUIPMENT AND CONSUMABLES REQUIRED
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ALIQUOTING PREPARED GENOMIC DNA To
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Negative Controls chapter 4 | 96-We
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Table 4.7 Nsp I Digestion Master Mi
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Stage 3: Ligation ABOUT THIS STAGE
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REAGENTS REQUIRED chapter 4 | 96-We
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DILUTE THE SAMPLES To dilute the sa
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Stage 4: PCR ABOUT THIS STAGE chapt
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About Controls chapter 4 | 96-Well
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B P1 P2 P3 Figure 4.2 Transferring
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Table 4.16 PCR Master Mix ADD PCR M
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WHAT YOU CAN DO NEXT chapter 4 | 96
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POOL THE PCR PRODUCTS chapter 4 | 9
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Figure 4.5 Loading Optical Plates w
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Process Control Metrics Evaluate th
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Table 4.24 PROBLEM: Average Sample
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IMPORTANT INFORMATION ABOUT THIS ST
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Figure 4.6 Work Area Set Up for Sta
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Stage 8: Labeling ABOUT THIS STAGE
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LOCATION AND DURATION • Main Lab
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Hybridizing Samples Using Heat Bloc
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Table 4.37 Hybridization Master Mix
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Load the Samples onto Arrays chapte
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Figure 4.8 Applying Tough-Spots ®
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Chapter 5 Washing, Staining, and Sc
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Introduction This chapter contains
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Reagent Preparation Wash A: Non-Str
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Experiment and Fluidics Station Set
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chapter 5 | Washing, Staining, and
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HANDLING THE GENECHIP ® PROBE ARRA
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Chapter 6 Fluidics Station Care and
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Introduction This chapter provides
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chapter 6 | Fluidics Station Care a
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White Clamp Figure 6.7 Releasing pe
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Figure 6.8 Troubleshooting decision
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PROBLEMS AND SOLUTIONS Table 6.3 Co
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Table 6.3 (Continued) Common error
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Table 6.4 (Continued) Common Error
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Table 6.5 (Continued) Other Problem
Page 191 and 192: Chapter 7 Analysis Workflow
Page 193 and 194: Introduction Software Requirements
Page 195 and 196: Figure 7.2 GTYPE main window with t
Page 197 and 198: Figure 7.3 Dynamic Model Mapping Al
Page 199 and 200: Table 7.1 Dynamic Model Mapping Alg
Page 201 and 202: FILE SETS chapter 7 | Analysis Work
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Page 205 and 206: NETAFFX SNP ANNOTATION chapter 7 |
Page 207 and 208: CALL RATE chapter 7 | Analysis Work
Page 209 and 210: chapter 7 | Analysis Workflow 197 G
Page 211 and 212: chapter 7 | Analysis Workflow 199 F
Page 213 and 214: chapter 7 | Analysis Workflow 201 S
Page 215 and 216: B2 OLIGO PERFORMANCE chapter 7 | An
Page 217 and 218: chapter 7 | Analysis Workflow 205 3
Page 219 and 220: chapter 7 | Analysis Workflow 207 W
Page 221 and 222: Chapter 8 Troubleshooting
Page 223 and 224: Assay Recommendations 211 Genotypin
Page 225 and 226: chapter 8 | Troubleshooting 213 10.
Page 227 and 228: Problem Likely Cause Solution Faint
Page 229 and 230: Problem Likely Cause Solution Posit
Page 231 and 232: Table 8.2 PROBLEM: Average Sample O
Page 233 and 234: When to Contact Technical Support c
Page 235 and 236: Appendix A Reagents, Equipment, and
Page 237 and 238: Reagents, Equipment, and Consumable
Page 239 and 240: appendix A | Reagents, Equipment, a
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Page 245 and 246: Consumables Required GENECHIP ® AR
Page 247 and 248: Supplier Contact List Table A.10 Su
Page 249 and 250: Appendix B Thermal Cycler Programs
Page 251 and 252: ABOUT THIS APPENDIX 500K DIGEST 500
Page 253 and 254: 500K FRAGMENT 500K LABEL 500K HYB a
Page 255 and 256: Appendix C Low Throughput Protocol
Page 257 and 258: Introduction The Affymetrix GeneChi
Page 259 and 260: BEFORE YOU BEGIN appendix C | Low T
Page 261 and 262: STEP 2: Restriction Enzyme Digestio
Page 263 and 264: DIGESTION PROCEDURE PRE-PCR CLEAN A
Page 265 and 266: STEP 3: Ligation REAGENTS AND EQUIP
Page 267 and 268: PCR STAGING AREA appendix C | Low T
Page 269 and 270: STEP 4: PCR REAGENTS AND EQUIPMENT
Page 271 and 272: PCR PROCEDURE PRE-PCR CLEAN ROOM ap
Page 273 and 274: MAIN LAB appendix C | Low Throughpu
Page 275 and 276: STEP 5: PCR Purification and Elutio
Page 277 and 278: appendix C | Low Throughput Protoco
Page 279 and 280: STEP 7: Fragmentation REAGENTS AND
Page 281 and 282: FRAGMENTATION PROCEDURE 1. Pre-heat
Page 287 and 288: LABELING PROCEDURE MAIN LAB appendi
Page 289 and 290: STEP 9: Target Hybridization REAGEN
Page 291 and 292: HYBRIDIZATION PROCEDURE appendix C
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appendix C | Low Throughput Protoco
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Appendix D Reagents, Instruments, a
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Appendix D
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appendix D | Reagents, Instruments,
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* Adaptor, Nsp 5' ATTATGAGCACGACAGA
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Table D.4 Reagents Supplied by Affy
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Fragmentation and Labeling appendix
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Mapping 250K/500K SNP assay

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