An Introduction to Molecular Biotechnology: Fundamentals, Methods and Applications

Molecular biotechnology continues to triumph, as this textbook testifies - edited by one of the academic pioneers in the field and written by experienced professionals. This completely revised second edition covers the entire spectrum, from the fundamentals of molecular and cell biology, via an overview of standard methods and technologies, the application of the various "-omics", and the development of novel drug targets, right up to the significance of system biology in biotechnology. The whole is rounded off by an introduction to industrial biotechnology as well as chapters on company foundation, patent law and marketing. The new edition features: - Large format and full color throughout - Proven structure according to basics, methods, main topics and economic perspectives - New sections on system biology, RNA interference, microscopic techniques, high throughput sequencing, laser applications, biocatalysis, current biomedical applications and drug approval - Optimized teaching with learning targets, a glossary containing around 800 entries, over 500 important abbreviations and further reading. The only resource for those who are seriously interested in the topic. Bonus material available online free of charge: www.wiley-vch.de/home/molecbiotech

Table of Contents:
Preface XIX List of Contributors XXI Abbreviations XXV Part I Fundamentals of Cellular and Molecular Biology 1 1 The Cell as the Basic Unit of Life 3 M. Wink 2 Structure and Function of Cellular Macromolecules 7 M. Wink 2.1 Structure and Function of Sugars 8 2.2 Structure of Membrane Lipids 10 2.3 Structure and Function of Proteins 14 2.4 Structure of Nucleotides and Nucleic Acids (DNA and RNA) 21 2.5 References 27 3 Structure and Functions of a Cell 29 M. Wink 3.1 Structure of a Eukaryotic Cell 29 3.2 Structure of Bacteria 50 3.3 Structure of Viruses 51 3.4 Differentiation of Cells 52 4 Biosynthesis and Function of Macromolecules (DNA, RNA, and Proteins) 57 M. Wink 4.1 Genomes, Chromosomes, and Replication 57 4.2 Transcription: From Gene to Protein 71 4.3 Protein Biosynthesis (Translation) 76 5 Distributing Proteins in the Cell (Protein Sorting) 81 M. Wink 5.1 Import and Export of Proteins via the Nuclear Pore 82 5.2 Import of Proteins in Mitochondria and Chloroplasts 83 5.3 Protein Transport into the Endoplasmic Reticulum 85 5.4 Vesicle Transport from the ER via the Golgi Apparatus to the Cytoplasmic Membrane 86 6 Evolution and Diversity of Organisms 91 M. Wink 6.1 Prokaryotes 91 6.2 Eukaryotes 91 Part II Standard Methods in Molecular Biotechnology 99 7 Isolation and Purification of Proteins 101 T. Wieland, M. Lutz 7.1 Introduction 101 7.2 Producing a Protein Extract 102 7.3 Gel Electrophoretic Separation Methods 103 7.4 Methods of Protein Precipitation 106 7.5 Column Chromatography Methods 107 7.6 Examples 113 8 Peptide and Protein Analysis with Electrospray Tandem Mass Spectrometry 115 A. Schlosser, W.D. Lehmann 8.1 Introduction 115 8.2 Principles of Mass Spectrometry 115 8.3 Mass Precision, Resolution, and Isotope Distribution 116 8.4 Principles of ESI 116 8.5 Tandem Mass Spectrometers 117 8.6 Peptide Sequencing with MS/MS 119 8.7 Identifying Proteins with MS/MS Data and Protein Databases 120 8.8 Determining Protein Molecular Mass 121 8.9 Analysis of Covalent Protein Modification 122 8.10 Relative and Absolute Quantification 123 9 Isolation of DNA and RNA 125 H. Weiher, R. Zwacka, I. Herr 9.1 Introduction 125 9.2 DNA Isolation 125 9.3 RNA Isolation 127 10 Chromatography and Electrophoresis of Nucleic Acids 129 H. Weiher, R. Zwacka, I. Herr 10.1 Introduction 129 10.2 Chromatographic Separation of Nucleic Acids 129 10.3 Electrophoresis 130 11 Hybridization of Nucleic Acids 133 H. Weiher, R. Zwacka, I. Herr 11.1 Significance of Base Pairing 133 11.2 Experimental Hybridization: Kinetic and Thermodynamic Control 133 11.3 Analytical Techniques 134 12 Use of Enzymes in the Modification of Nucleic Acids 137 A. Groth, R. Zwacka, H. Weiher, I. Herr 12.1 Restriction Enzymes (Restriction Endonucleases) 137 12.2 Ligases 139 12.3 Methyltransferases 139 12.4 DNA Polymerases 140 12.5 RNA Polymerases and Reverse Transcriptase 141 12.6 Nucleases 141 12.7 T4 Polynucleotide Kinase 141 12.8 Phosphatases 142 13 Polymerase Chain Reaction 143 A. Mohr, H. Weiher, I. Herr, R. Zwacka 13.1 Introduction 143 13.2 Techniques 143 13.3 Areas of Application 146 14 DNA Sequencing 149 R. Zwacka, A. Mohr, I. Herr, H. Weiher 14.1 Introduction 149 14.2 DNA Sequencing Methods 149 14.3 Strategies for Sequencing the Human Genome 151 14.4 Practical Significance of DNA 152 15 Cloning Procedures 153 T. Wieland, S. Lutz 15.1 Introduction 153 15.2 Construction of Recombinant Vectors 153 16 Expression of Recombinant Proteins 169 T. Wieland, S. Lutz 16.1 Introduction 169 16.2 Expression of Recombinant Proteins in Host Organisms 170 16.3 Expression in Cell-Free Systems 178 17 Patch Clamp Method 181 R. Kraft 17.1 Biological Membranes and Ion Channels 181 17.2 Physical Foundations of the Patch Clamp Method 182 17.3 Patch Clamp Configurations 182 17.4 Applications of the Patch Clamp Method 184 18 Cell Cycle Analysis 187 S. Wölfl, A. Kitanovic 18.1 Analyzing the Cell Cycle 187 18.2 Experimental Analysis of the Cell Cycle 189 19 Microscopic Techniques 197 S. Diekmann 19.1 Electron Microscopy 197 19.2 Atomic or Scanning Force Microscopy 200 19.3 Light Microscopy 202 19.4 Microscopy in the Living Cell 206 20 Laser Applications 211 M. Vogel, R. Fink 20.1 Principles of Laser Technology 211 20.2 Properties of Laser Radiation 213 20.3 Types of Lasers and Setups 213 20.4 Applications 214 Part III Key Topics 217 21 Genomics and Functional Genomics 219 S.Wiemann, M. Frohme 21.1 Introduction 219 21.2 Technological Developments in DNA Sequencing 221 21.3 Genome Sequencing 222 21.4 cDNA Projects 238 21.5 Functional Genomics 246 21.6 Identification and Analysis of Individual Genes 248 21.7 Investigation of Transcriptional Activity 253 21.8 Cell-based Methods 266 21.9 Functional Analysis of Entire Genomes 272 22 Bioinformatics 275 B. Brors, K. Fellenberg 22.1 Introduction 275 22.2 Data Sources 276 22.3 Sequence Analysis 279 22.4 Evolutionary Bioinformatics 283 22.5 Gene Prediction 287 22.6 Bioinformatics in Transcriptome and Proteome Analysis 288 22.7 Bioinformatic Software 293 23 Cellular Systems Biology 295 H. Schmidt-Gienewinkel, S. Legewie, B. Brors, R. König 23.1 Introduction 295 23.2 Analysis of Cellular Networks by Top-Down Approaches 296 23.3 Overview of Bottom-Up Modeling of Biochemical Networks 304 23.4 Biological Examples 309 24 Protein–Protein and Protein–DNA Interaction 315 P. Uetz, E. Pohl 24.1 Protein–Protein Interactions 315 24.2 Protein–DNA Interactions 324 25 Drug Research 331 M. Koegl, R. Tolle, U. Deuschle, C. Kremoser 25.1 Introduction 331 25.2 Active Compounds and their Targets 331 25.3 Preclinical Pharmacology and Toxicology 344 25.4 Clinical Development 346 25.5 Clinical Testing 346 26 Drug Targeting and Prodrugs 349 G. Fricker 26.1 Drug Targeting 349 26.2 Prodrugs 355 26.3 Penetration of Drugs through Biological Membranes 356 26.4 Prodrugs to Extend Duration of Effect 357 26.5 Prodrugs for the Targeted Release of a Drug 357 26.6 Prodrugs to Minimize Side Effects 358 27 Molecular Diagnostics in Medicine 359 S. Wölfl, R. Gessner 27.1 Uses of Molecular Diagnostics 359 27.2 Which Molecular Variations Should be Detected 364 27.3 Molecular Diagnostic Methods 367 27.4 Outlook 375 28 Recombinant Antibodies and Phage Display 377 S. Dübel 28.1 Introduction 377 28.2 Why Recombinant Antibodies? 379 28.2.1 Recombinant Antibodies are Available In Vitro without Immunization 379 28.3 Obtaining Specific Recombinant Antibodies 379 28.4 Production of Recombinant Antibodies 384 28.5 Formats for Recombinant Antibodies 386 28.6 Applications of Recombinant Antibodies 392 28.7 Outlook 394 29 Transgenic and Gene-Targeted Mice and their Impact in Medical Research 395 R. Sprengel 29.1 Overview 395 29.2 Transgenic Mice 395 29.3 Homologous Recombination: knock-out (-in) mice 398 29.4 Conditionally Regulated Gene Expression 399 29.5 Impact of Genetically Modified Mice in Biomedicine 400 29.6 Outlook 402 30 Gene Therapy: Strategies and Vectors 403 A. Groth, I. Herr 30.1 Introduction 403 30.2 Principles of Somatic Gene Therapy 404 30.3 Germ Line Therapy 405 30.4 Setbacks in Gene Therapy 406 30.5 Vectors for Gene Therapy 406 30.6 Specific Expression 413 31 RNA Interference, Modified DNA, Peptide Nucleic Acid, and Applications in Medicine and Biotechnology 415 N. Metzler-Nolte, A. Sosniak 31.1 Introduction 415 31.2 Modified Nucleic Acids 416 31.3 Interactions of DNA Analogs with Complementary DNA and RNA 419 31.4 RNAi 421 31.5 Applications 424 32 Plant Biotechnology 431 H. Hillebrand, R. Hell 32.1 Introduction 431 32.2 Gene Expression Control 433 32.3 Production of Transgenic Plants 434 32.4 Selection of Transformed Plant Cells 439 32.5 Regeneration of Transgenic Plants 445 32.6 Plant Analysis: Identification and Characterization of Genetically Engineered Plants 446 33 Biocatalysis in the Chemical Industry 451 M. Breuer, B. Hauer 33.1 Introduction 451 33.2 Bioconversion/Enzymatic Procedures 454 33.3 Development of an Enzyme for Industrial Biocatalysis 456 33.4 Fermentative Procedures 462 Part IV Biotechnology in Industry 473 34 Industrial Application: Biotech Industry, Markets, and Opportunities 475 J. Schüler 34.1 Historical Overview and Definitions of Concepts 475 34.2 Areas of Industrial Application of Molecular Biotechnology 476 34.3 Status Quo of the Biotech Industry World-Wide 485 35 Patents in the Molecular Biotechnology Industry: Legal and Ethical Issues 487 David B. Resnik 35.1 Patent Law 487 35.2 Ethical and Policy Issues in Biotechnology Patents 492 35.3 Conclusions 498 36 Drug Approval in the European Union and United States 499 G. Walsh 36.1 Introduction 499 36.2 Regulation within the European Union 499 36.3 Regulation in the United States 503 36.4 Advent and Regulation of Biosimilars 506 36.5 International Regulatory Harmonization 506 37 Emergence of a Biotechnology Industry 509 C. Kremoser 38 The 101 of Founding a Biotech Company 517 C. Kremoser 38.1 First Steps Towards Your Own Company 517 38.2 Employees: Recruitment, Remuneration, Participation 522 39 Marketing 527 C. Kremoser 39.1 Introduction 527 39.2 What Types of Deals are Possible? 528 39.3 What Milestone or License Fees are Effectively Paid in a Biotech/Pharma Cooperation? 529 39.4 PR and IR in Biotech Companies 530 Appendix 533 Further Reading 535 Glossary 551 M. Wink Subject Index 587