Úplné zobrazení záznamu

Toto je statický export z katalogu ze dne 21.05.2022. Zobrazit aktuální podobu v katalogu.

Bibliografická citace

.
0 (hodnocen0 x )
EB
EB
ONLINE
5th edition
London : Academic Press, [2017]
1 online zdroj : ilustrace
Externí odkaz    Plný text PDF 
   * Návod pro vzdálený přístup 


ISBN 9780128046791 (e-kniha)
ISBN 9780128046784 (print)
Obsahuje bibliografické odkazy a rejstřík
Popsáno podle tištěné verze
001483094
Preface xvii // 1. RNA and the Cellular Biochemistry Revisited // Why Study RNA? 1 // What is RNA? 2 // Polynucleotide Synthesis 5 // Types of RNA 9 // Transcription and the Central Dogma 11 // Promoters, Transcription Factors, and Regulatory Elements 13 // Gene and Genome Organization Affect Transcription 18 // RNA Polymerases and the Products of Transcription 23 // Hallmarks of a Typical mRNA 27 // 5’ Cap 29 // 5’ UTR (Leader Sequence) 30 // Coding Region 31 // 3’ DTR (Trailer Sequence) 32 // Poly(A) Tail 32 // Organellar mRNAs 34 // mRNA Stability and Turnover 35 // Bicistronic mRNAs 38 // Prokaryotic mRNAs 38 // mRNA Sequence and Structure Affect Translation 39 // Levels of Gene Regulation 41 // Alternative Splicing of mRNA from a Single Genetic Locus 43 // Trans-Splicing: mRNA Repair 44 // Overview of Noncoding RNAs 46 // References 47 // Further Reading 53 // 2. Creating a Ribonuclease-Free Environment // Rationale 55 // Elimination of Resilient Ribonucleases 56 // Latent RNase Contamination Issues 58 // Types of Ribonuclease Inhibitors 59 // Specific Inhibitors 60 // Nonspecific Inhibitors 62 // v // vi Contents // Preparation of Equipment and Reagents 63 // Diethyl Pyrocarbonate (DEPC) 64 // Alternative: USP Sterile Water 66 // Hydrogen Peroxide 66 // NaOH and SDS 67 // Other Reagents Used to Control Nuclease Activity 67 // Guanidinium Salts 67 // Sodium Dodecyl Sulfate 68 // N-Laurylsarcosine 68 // Phenol:Chloroform:lsoamyl Alcohol 68 // 8-Hydroxyquinoline 69 // Cesium Chloride 70 // Cesium Trifluoroacetate 70 // Proteinase ? 70 // RNA/arer 71 // Protocol: Synthesis of VDR 71 // References 73 // 3. RNA Isolation Strategies // Rationale 75 // Goals in the Purification of RNA 77 // Lysis Buffer Formulations 80 // Gentle Lysis Buffers 80 // Chaotropic Lysis Buffers 87 // Isolation of RNA with Guanidinium Buffers 88 // Guanidinium-Acid-Phenol Extraction Techniques 89 //
Protocol: Guanidinium-Acid-Phenol Extraction 91 // Density Gradient Centrifugation 92 // Cesium Chloride 93 // Cesium Trifluoracetate (CsTFA) 97 // Isolation of RNA and DNA from the Same Source 101 // Protocol: Simultaneous Isolation of RNA and DNA 101 // The Word on Kits 104 // Silica Technology 105 // Isolation of Cytoplasmic RNA on a Silica Column 106 // Affinity Matrices 107 // Other Methods 107 // Protocol: Rapid Isolation of RNA with SDS and // Potassium Acetate Reagents 108 // Protocol: Isolation of Prokaryotic RNA 109 // Protocol: Isolation of RNA From Yeast 110 // Short- and Long-Term Storage of Purified RNA 112 // References 114 // 4. The Truth About Tissues // Rationale -j -j 7 // Tissue Culture or Tissue? 117 // Contents vii // Advantages of Cell Culture 118 // Advantages of Tissue Samples 119 // Homogenization Methods 120 // Motorized Homogenizers 120 // ?ounce Homogenization 121 // BeadBeater Homogenization 123 // RNA Isolation Strategies for Various Organs and Tissues 125 // Fresh Tissue 126 // Frozen Tissue 127 // Fixed Tissue 128 // Protocol: LiCI-Urea Method for RNA Isolation From Tissue 130 // Protocol: RNA Isolation From Lipid-Enriched Samples 133 // Purification of Polysome- and Protein-Engaged mRNA 135 // Protocol: Isolation of Polysomal mRNA 137 // Collecting Samples in the Field 139 // RNA "Clean-Up" Methods 140 // Troubleshooting RNA Isolation From Tissue 142 // References 143 // Further Reading 144 // 5. Going Green: RNA and the Molecular Biology of Plants // Rationale 145 // RNA Isolation and the Peculiarities of Plants 145 // Types of RNA Produced in Plant Cells 149 // Protocol: RNA Isolation From Leaf 151 // Protocol: RNA Isolation From Bark 153 // Protocol: RNA Isolation From Fruit 155 // Protocol: RNA Isolation From Plant Tissue With Hot Borate 158 // Strategies for RNA Isolation From Other Plant Tissues 160 //
Troubleshooting RNA Isolation From Plant Tissue 161 // References 162 // Further Reading 164 // 6. Quality Control for RNA Preparations // Rationale 167 // Quality Control Technique 1: UV Spectrophotometry // and Absorption Ratios 167 // Determination of Nucleic Acid Concentration 168 // Determination of Nucleic Acid Purity 171 // Nonspectrophotometric Methods 174 // Quality Control Technique 2: Electrophoretic Profiling of RNA 176 // Protocol: Nondenaturing Agarose Electrophoresis 179 // Quality Control Technique 3: RNA Integrity Number (RIN) 180 // Quality Control Technique 4: UV Shadowing 182 // Protocol: UV Shadowing 182 // Quality Control Technique 5: Sample Capacity to Support RT-PCR 184 // Quality Control Technique 6: Sample Capacity to // Support Northern Analysis 184 // Quality Control Technique 7: Sample Capacity to // Support In Vitro Translation 185 // References 185 // 7. cDNA-A Permanent Biochemical Record of the Cell // Rationale 187 // cDNA Synthesis-An Overview 187 // First-Strand Considerations 189 // Reverse Transcriptase Options 193 // Second-Strand Considerations 197 // Classical Methods 198 // PCR-Based Methods 199 // Protocol: First-Strand cDNA Synthesis 199 // Assessing cDNA Synthesis Efficiency 201 // Cloning cDNA 202 // Ligation Considerations 204 // Enzymes Used for Ligation 205 // Applications 206 // References 206 // 8. RT-PCR: A Science and an Art Form // Rationale 209 // PCR-An Overview 209 // RT-PCR-General Approach 217 // PCR Carryover Prevention 219 // Laboratory Design 220 // Procedural Methods 221 // Aerosol-Resistant Tips 222 // U raci I-N-Glycosy lase 223 // Primer Design 223 // Basic Rules 228 // Tm Considerations 233 // AG Considerations 234 // Multiplex Primer Design 235 // Optimization Procedures 235 // Thermostable Polymerases 240 // Positive Controls 241 // Negative Controls 241 // Hot-Start PCR 242 // Locked Nucleic Acid 244 //
Touchdown PCR 246 // Internal Controls 246 // The Word on Transcription Controls 249 // Analysis of PCR Products 250 // RT-PCR Quality Control Points 251 // Non-PCR Methods for Confirming PCR-Derived Data 254 // Related Techniques 255 // 5’ RACE PCR 255 // 5’ RLM-RACE 256 // 3’ RACE PCR 258 // Nested PCR 259 // Long-Range PCR 260 // Single-Cell PCR 263 // Splinkerette PCR 264 // The Hunt for Alternative Transcription Start Sites 265 // Protocol: First-Strand cDNA Synthesis 266 // Protocol: PCR Amplification of cDNA 267 // Cloning PCR Products 268 // Protocol: ?-Tailing of Blunt-End PCR Products 270 // Protocol: ?? Cloning Ligation Reaction 270 // TOPO Cloning 272 // Other Amplification Procedures 272 // Linear RNA Amplification (Eberwine Process) 273 // Strand Displacement Amplification 274 // Nucleic Acid Sequence Based Amplification (NASBA) 275 // Rolling Circle Amplification (RCA) 275 // Ligase Chain Reaction 275 // LAMP 276 // References and Suggested Reading 276 // 9. Quantitative PCR Techniques // Rationale 283 // Sensitivity Index 284 // Quantitative Approaches 285 // The MIQE Guidelines 287 // Real-time PCR 288 // Real-Time PCR Platforms 292 // SYBR Green Assay 293 // TaqMan Assay 294 // Molecular Beacons 295 // Scorpions 297 // Melting Curve Analysis 298 // Digital PCR 300 // Internal Controls 302 // Exogenous Controls 302 // Control Reaction Formats 304 // Negative Control Considerations 308 // PCR Arrays 310 // Competitive PCR: Key Considerations 311 // Competitive PCR: Major Steps Involved 316 // Alternative Approach: Nonreal-Time Competitive PCR 318 // Protocol: Competitive PCR 318 // Troubleshooting Quantitative PCR Techniques 324 // References 327 // 10. miRNA // Rationale 329 // Structural and Functional Characteristics of miRNA 329 // miRNA Biogenesis 331 // Differences and Similarities of miRNA and siRNA 335 // miRNA Profiling 336 //
miRNA as Key Regulator of Gene Expression 338 // miRNA Biomarkers 340 // References and Suggested Reading 342 // 11. RNA Interference and RNA Editing // Rationale 345 // Essential RNAi Terminology 346 // RNA Interference -How It Works 348 // Endogenous Silencing Pathways 351 // Exogenous Silencing Strategies 353 // Effective Design of siRNAs 359 // RNAi and Alternative Transcript Splicing 363 // In Vitro and In Vivo Issues 363 // RNAi Validation 367 // RT-PCR Approaches 367 // Northern Analysis 367 // Western Analysis and Other Protein Methods 368 // RNAi Applications 368 // CRISPR-Cas9 369 // References 373 // Further Reading 376 // 12. Stringency: Conditions That Influence Nucleic Acid Structure // Rationale 377 // Types of Double-Stranded Molecules 377 // Importance of Controlling Stringency 378 // Effect of Salt on Stringency 380 // Effect of pH on Stringency 380 // Effect of Temperature on Stringency 381 // Effect of Formamide on Stringency 381 // Effect of Urea on Stringency 382 // References 382 // 13. Electrophoresis of RNA // Rationale 383 // Normalization of Samples by Nucleic Acid Concentration 384 // Direct Measurement of Poly(A) Content 385 // Protocol: Poly(A) Normalization With a Poly(T) Probe 387 // Intramolecular Base-Pairing Mandates RNA Denaturation 390 // Formaldehyde Denaturation 392 // Protocol: Formaldehyde-Denaturing Gels 393 // Urea Denaturation 395 // Protocol: Urea Denaturation 395 // Glyoxal/Dimethyl Sulfoxide Denaturation 396 // Protocol: Glyoxalation and Electrophoresis of RNA 397 // Gel and Sample Preparation 398 // Running RNA on Nondenaturing Gels 399 // Molecular Weight Standards 400 // Proper Use of Size Standards 401 // Ribosomal RNA 402 // Gel Staining Options 410 // Ethidium Bromide 410 // SYBR Green 413 // SYBR Gold 413 // SYBR Safe 415 // GelStar 416 // Silver Staining 416 // Acridine Orange 416 // Methylene Blue 417 //
Safety Considerations and Equipment Maintenance 418 // A Few Tips for Running Agarose Gels for the First Time 420 // References 424 // 14. Photodocumentation and Image Analysis // Rationale 427 // Safety First 427 // Digital Image Analysis 428 // Image Enhancement 431 // Filtration 434 // Image Formats 436 // Practical Considerations 436 // Image Analysis Training 439 // Phosphorlmagers 440 // Traditional Methods of Photodocumentation 440 // Camera Settings 441 // Tips for Optimizing Electrophoretograms 442 // Inherent Limitations of Photographic and X-ray Films 445 // References and Suggested Reading 446 // 15. Northern Analysis // Rationale 447 // Choice of Blotting Membrane 448 // Nitrocellulose 449 // Nylon 450 // Polyvinylidene Difluoride 451 // Handling and Membrane Preparation 451 // Northern Transfer Techniques 452 // Capillary Transfer 452 // TurboBlotter 454 // Vacuum Blotting 454 // Electroblotting 455 // Alkaline Blotting 456 // Protocol: RNA Transfer by Passive Capillary Diffusion 457 // Protocol: Turboblotter Downward Transfer of RNA 459 // Immobilization Techniques 462 // Baking 463 // Crosslinking by UV Irradiation 463 // Protocol: UV Crosslinking RNA to Nylon Filters 464 // Postfixation Handling of Blotting Membranes 465 // Reverse Northern Analysis 466 // References 467 // 16. Nucleic Acid Probe Technology // Rationale 469 // DNA Probes 470 // DNA Probe Synthesis 471 // Sense and Antisense RNA Probes 475 // RNA Probe Synthesis 477 // Selection of Labeling System 479 // Isotope Labeling 479 // Popular Chemiluminescence Platforms 482 // Direct Enzyme Labeling 483 // Biotin 484 // Digoxigenin 486 // Fluorescein 487 // The Ubiquitous Dyes Cy3 and Cy5 487 // Probe Purification 488 // Probe Storage 489 // Factors Influencing Hybridization Kinetics and Duplex Stability 490 // Temperature and Tm Considerations 491 // Ionic Strength 494 // pH 494 //
Probe Length 494 // Probe Concentration 494 // G + C Content 495 // Contents xiii // Mismatching 496 // Probe Complexity 497 // Example 497 // Viscosity 497 // Formamide 498 // Urea 498 // Mixed-Phase Hybridization: Northern and Southern Blots 499 // Prehybridization: Filter Preparation 499 // Protocol: Prehybridization for Long Probes 500 // Protocol: Generic Method for Probe Removal 503 // Principles of Detection 505 // Digital Imaging Systems 505 // Autoradiography 506 // Nonisotopic Procedures 517 // Biotin 518 // Digoxigenin 518 // Fluorescein and Fluorescence Imaging 519 // Direct Enzyme Labeling 519 // Detection by Chemiluminescence 519 // Substrates for Chemiluminescence 521 // Chromogenic Detection Procedures 523 // References 524 // 17. Isolation of Polyadenylated RNA // Rationale 527 // Polyadenylation 528 // Selection of Polyadenylated Molecules: How It Works 529 // The Poly(A) Caveat 531 // Example 1 - cDNA Synthesis Considerations 531 // Example 2 - Assay Sensitivity Considerations 532 // Magnetic Bead Technology for Poly(A)+ Purification 533 // Oligo(dT)-Cellulose Chromatography 535 // Protocol: Noncolumn Poly(A)+ Purification 536 // Protocol: Large-Scale PoIy(A)+ RNA Purification with // Oligo(dT)-CelIulose Beads 538 // References 543 // 18. Quantification of Specific mRNAs by Nuclease Protection // Rationale 545 // Basic Approach 546 // Probe Selection 551 // Optimization Suggestions 555 // Potential Difficulties 556 // Protocol: Transcript Quantification by S1 Analysis 558 // xiv Contents // Protocol: Transcript Quantification by RNase Protection 562 // Troubleshooting 565 // References 566 // 19. Analysis of Nuclear RNA // Rationale 569 // Transcription Rate Assays 569 // Relationship to the Study of Steady-State RNA 574 // Nuclear Run-off Versus Nuclear Run-on Assay 576 // Protocol: Nuclear Run-on Assay 577 // Harvesting of Cells and Preparation on Nuclei 577 //
Alternative Protocol for Preparation of Nuclei From Cell Culture 578 // Alternative Protocol for Preparation of Nuclei From Whole Tissue 580 // Labeling and Recovery of Transcripts 581 // Preparation of Target DNA 583 // Preparation of RNA for Hybridization 585 // Posthybridization Washes and Detection 586 // Protocol: Alternative Procedure for Nuclear Run-on Assay 587 // Protocol: Nuclease Protection-Pulse Label Transcription Assay 588 // Distinguishing Among the Activities of RNA Polymerases 591 // Extraction of Nuclear RNA for Steady-State Analysis 592 // Protocol: Direct Isolation of Nuclear RNA 593 // Protocol: Preparation of Nuclear RNA From Cells Enriched in // Ribonuclease 595 // Troubleshooting Nuclear RNA Analysis 596 // References 597 // Further Reading 599 // 20. Nonarray Methods for Global Analysis of Gene Expression // Rationale 601 // Essential Issues 602 // Subtractive Methods 603 // Suppression Subtractive Hybridization (SSH) 603 // Troubleshooting 611 // Nonsubtractive Methods 613 // mRNA Differential Display 614 // Troubleshooting 623 // References 625 // 21. Genomes, Transcriptomes, Proteomes, and Bioinformatics // Rationale 629 // Essential Nomenclature 631 // Transcriptomes and Transcriptomics 632 // RNA-Seq: The Premier Transcriptomics Tool 633 // Contents XV // Genomes and Genomics 639 // Proteomes and Proteomics 641 // RNA-RNA and RNA-Protein Interactions 646 // Bioinformatics 648 // Search for Genes -Have a BLAST! 649 // References 652 // 22. RNA Biomarker Discovery // Rationale 655 // Biomarkers Defined 655 // Characteristics of Useful Biomarkers 657 // Circulating RNA 658 // Identification of Biomarkers for Research and Diagnostic // Applications 659 // DNA Approaches 660 // RNA Approaches 661 // Protein Approaches 663 // Metabolomics Approaches 664 // Biomarker Issues and Shortcomings 664 // References and Suggested Reading 665 //
23. High-Throughput Analysis of Gene Expression // Rationale 667 // What is a Microarray? 667 // What Is a Heat Map? 672 // What Microarrays Can Do 674 // What Microarrays Cannot Do 674 // Major Steps in Microarray Analysis 677 // Reference RNA 679 // What Is a Macroarray? 680 // Applications 682 // References 684 // 24. Functional Genomics and Transcript Profiling // Rationale 685 // Functional Genomics Defined 686 // Importance of Functional Genomics Approaches 686 // Commonly Used Functional Genomics Approaches 687 // Relationship of Functional Genomics Approaches // to Classical Molecular Biology 693 // References 695 // Further Reading 695 // xvi Contents // 25. A Few RNA Success Stories // A Typical Experiment? 698 // Sensitivity Issues 701 // What to Do Next 701 // Where to Turn for Help 703 // Current Innovations 704 // RNA Biobanking 704 // RNA Reprogramming 705 // References 706 // Epilogue: A Few Pearls of Wisdom 707 // Appendix A: Maintaining Complete and Accurate Records 715 // Appendix B: Converting Mass to Moles 719 // Appendix C: Disposal of Ethidium Bromide and SYBR Green Solutions 723 // Appendix D: Removal of DNA From an RNA Sample 727 // Appendix E: Removal of RNA From a DNA Sample 729 // Appendix F: Useful Stock Solutions for the Molecular Biologist 731 // Appendix G: Silanizing Centrifuge Tubes and Glassware 737 // Appendix H: Dot Blot Analysis 739 // Appendix I: Electrophoresis: Principles, Parameters, and Safety 749 // Appendix J: Polyacrylamide Gel Electrophoresis 761 // Appendix K: Centrifugation as a Mainstream Tool for the Molecular // Biologist 765 // Appendix L: Trypsinization Protocol for Anchorage-Dependent Cells 773 // Appendix M: Isolation of High-Molecular-Weight DNA by Salting-Out 777 // Appendix N: Phenol Preparation 781 // Appendix O: Deionization of Formamide, Formaldehyde, and Glyoxal 785 //
Appendix P: Selected Suppliers of Equipment, Reagents, and Services 787 // Appendix Q: Useful SI Prefixes and Common SI Base Units 797 // Appendix R: Common Abbreviations 799 // Appendix S: Trademark Citations 801 // Glossary 805 // Index 833
(OCoLC)1001274603
GBB7J0382

Zvolte formát: Standardní formát Katalogizační záznam Zkrácený záznam S textovými návěštími S kódy polí MARC