Molecular Biotechnology (5^th Ed.) Principles and Applications of Recombinant DNA ASM Books Series

Since 1994, Molecular Biotechnology: Principles and Applications of Recombinant DNA has introduced students to the fast-changing world of molecular biotechnology. With each revision, the authors have extensively updated the book to keep pace with the many new techniques in gene isolation and amplification, nucleic acid synthesis and sequencing, gene editing, and their applications to biotechnology. In this edition, authors Bernard R. Glick and Cheryl L. Patten have continued that tradition, but have also overhauled the book?s organization to

• Detail fundamental molecular biology methods and recombinant protein engineering techniques, which provides students with a solid scientific basis for the rest of the book.
• Present the processes of molecular biotechnology and its successes in medicine, bioremediation, raw material production, biofuels, and agriculture.
• Examine the intersection of molecular biotechnology and society, including regulation, patents, and controversies around genetically modified products.

Filled with engaging figures that strongly support the explanations in the text, Molecular Biotechnology: Principles and Applications of Recombinant DNA presents difficult scientific concepts and technically challenging methods in clear, crisp prose.

This excellent textbook is ideal for undergraduate and graduate courses in introductory biotechnology, as well as, courses dedicated to medical, agricultural, environmental, and industrial biotechnology applications.

Preparation of DNA for Cloning 11

Insertion of Target DNA into a Plasmid Vector 16

Transformation and Selection of Cloned DNA in a Bacterial Host 20

Cloning Eukaryotic Genes 24

Recombinational Cloning 28

Genomic Libraries 30

Amplification of DNA by PCR 36

Cloning PCR Products 39

Quantitative PCR 39

Synthesis of Oligonucleotides 42

Assembling Oligonucleotides into Genes 48

Gene Synthesis by PCR 50

Dideoxynucleotide Procedure 53

Pyrosequencing 55

Sequencing using Reversible Chain Terminators 57

Sequencing by Single Molecule Synthesis 57

Preparation of Genomic DNA Sequencing Libraries 60

High-Throughput Next-Generation Sequencing Strategies 61

Genome Sequence Assembly 63

Sequencing Metagenomes 64

Transcriptomics 67

Proteomics 72

Metabolomics 85

Regulation of Transcription 94

Increasing Translation Efficiency 98

Increasing Protein Stability 102

Increasing Protein Secretion 106

Facilitating Protein Purification 110

DNA Integration into the Host Chromosome 115

Posttranslational Modification of Eukaryotic Proteins 120

General

Features of Eukaryotic Expression Systems 122

Yeast Expression Systems 124

Baculovirus–Insect Cell Expression Systems 136

Mammalian Cell Expression Systems 143

Directed Mutagenesis 154

Random Mutagenesis 158

Examples of Protein Engineering 162

Antibodies 178

Agglutination 183

Enzyme-Linked Immunosorbent Assays 183

Protein Arrays to Detect Polygenic Diseases 189

Immunoassays for Protein Conformation-Specific Disorders 191

Hybridization Probes 193

PCR-Based Detection Methods 200

DNA Microarrays 208

Whole Genome Sequencing to Assess Genetic Disease Risk 214

Detecting RNA Signatures of Disease 215

Detection of Disease-Associated Changes in Gene Expression

Using Microarrays 215

Detection of RNA Signatures of Antibiotic Resistance in Bacteria 216

Detection of miRNA Signatures of Cancers 217

Fluorescent Proteins

219

Luciferase 221

Microbial Biosensors 222

Human Interferons 231

Human Growth Hormone 235

Tumor Necrosis Factor Alpha 237

Extending Protein Half-Life 238

DNase

I 240

Alginate Lyase 242

Phenylalanine Ammonia Lyase 245

α1-Antitrypsin 247

Glycosidases 248

Masking Nonhuman Epitopes 249

Engineering Bacteriophages 250

Targeting Mitochondria 253

Interleukin-10 255

Leptin 258

An HIV Inhibitor 258

Insulin 260

Hybrid Human–Mouse Monoclonal Antibodies 264

Human Monoclonal Antibodies 268

Antibody Fragments 270

Combinatorial Libraries of Antibody Fragments 274

A Combinatorial Library of Full-Length Antibodies 277

Shuffling CDR Sequences 278

Dual-Variable-Domain Antibodies 280

Anticancer Antibodies 281

Antibodies Against Various Diseases 284

Antiobesity Antibodies 287

Enhanced Antibody Half-Life 290

Nucleic Acids as Therapeutic Agents 297

Targeting Specific mRNA and DNA Sequences

299

Antisense RNA 299

Aptamers 302

Ribozymes and DNAzymes 307

Interfering RNA 311

Zinc Finger Nucleases 315

CRISPR-Cas System 317

Nanozymes 318

Nanoparticles 319

Direct Injection 325

Lipids 327

Bacteria 328

Dendrimers 331

Antibodies 332

Aptamers 332

Transposons 334

Prodrug Activation Therapy 335

Promoterless Gene Targeting 337

Herpes Simplex Virus 348

Cholera 350

SARS 350

Staphylococcus aureus

351

Human Papillomavirus 353

Foot-and-Mouth Virus 354

Streptococcus 356

Delivery 357

Malaria 359

Delivery 364

Cancer 370

Dental Caries 370

Herpes Simplex Virus 372

Cholera 374

Salmonella Species 375

Leishmania Species 378

Vaccines Directed against Viruses 378

Vaccines Directed against Bacteria 388

Bacteria as Antigen Delivery Systems 392

Influenza Virus 396

L-Ascorbic Acid 407

Indigo 410

Amino Acids 412

Lycopene 417

Antibiotics 418

Biopolymers 429

Genetic Engineering of Biodegradative Pathways 436

Commercial Production of Fructose and Alcohol 446

Increasing Alcohol Production 448

Improving Fructose Production 453

Lignocellulosics 455

Cellulase Genes 457

Direct Conversion of Biomass to Ethanol 462

Zymomonas mobilis 464

Batch Fermentation 477

Fed-Batch Fermentation 479

Continuous Fermentation 480

Maximizing The Efficiency of The Fermentation Process 481

High-Density Cell Cultures 483

Increasing Plasmid Stability 484

Quiescent E. Coli Cells 485

Protein Secretion 486

Reducing Acetate 489

Two-Stage Fermentation in Tandem Airlift Reactors 495

Two-Stage Fermentation in a Single Stirred-Tank Reactor 496

Batch versus

Fed-Batch Fermentation 498

Protein Solubilization 506

Utilizing an Immobilized Enzyme 507

Magnetic Separation of Proteins 507

Very High Level Protein Expression 529

Transient Gene Expression 533

Plant Promoters 536

Targeted Gene Editing 538

Facilitating Protein Purification 539

Protein Glycosylation 541

Removing Marker Genes from Nuclear DNA 543

Removing Marker Genes from Chloroplast DNA 545

11 Transgenic Plants 551

Insect Resistance 551

B. thuringiensis Insecticidal Toxin 551

Increasing Expression of the B. thuringiensis Protoxin 555

Other Strategies for Protecting Plants against Insects 558

Preventing the Development of B. thuringiensis-Resistant

Insects 564

Targeting Aphids 569

Viral Coat Protein-Mediated Protection 570

Protection by Expression of Other Genes 574

Amino Acids 594

Lipids 595

Vitamins 599

Iron 601

Gluten 602

Preventing Discoloration 603

Starch 605

Antibodies 608

Poly(3-hydroxybutyric Acid) 610

Altering Lignin Content 615

Increasing Oxygen Content 618

DNA Microinjection Method 627

Retroviral Vector Method 629

Engineered Embryonic Stem Cell Method 631

Conditional Gene Inactivation with the Cre–loxP

Recombination System 637

Genome Editing with the CRISPR-Cas System

641

Gene Knockdown by RNA Interference 643

Transgenic Mice: Applications 644

Transgenic Disease Models: Alzheimer Disease 644

Transgenic Mice as Test Systems 647

Control of Transgene Expression 651

Conditional Control of Cell Death 654

Cloning Livestock by Somatic Cell Nuclear Transfer 656

Production of Pharmaceuticals 658

Production of Donor Organs 660

Disease Resistant Livestock 661

Improving Milk Quality 664

Improving Animal Production Traits 665

Environmental Concerns 682

Regulations 683

Food Ingredients Produced by Genetically Engineered

Microorganisms 684

Genetically Modified Crops 687

Genetically Engineered Livestock 691

Alteration of Nutritional Content of Food 692

Potential for Introducing Toxins or Allergens into Food 696

Potential for Transferring Transgenes from Food to Humans

or Intestinal Microorganisms 698

Controversy About the Labeling of Genetically Modified Foods 700

Impact of Genetically Engineered Crops on Biodiversity 700

Who Benefits from Production of Genetically Modified Foods? 703

Environmental Benefits of Genetically Modified Crops 704

How do Views about Genetically Engineered Organisms

Impact Trade? 705

New Biological Drugs 706

Genetic and Genomic Testing 709

Economic Issues 711

Patenting Biotechnology 714

Patenting 714

Patenting in Different Countries 716

Patenting Nucleic Acid Sequences 717

Patenting Living Organisms 719

Patenting and Fundamental Research 720