Genetic Modification of Plants (eBook)

Part A: Generation and Analysis of Transgenic Plants 27
Chapter 1: Plant Nuclear Transformation 28
Introduction to Plant Transformation 28
DNA Introduction Basics 28
Transient Expression 29
Optimization of Transient Expression 30
Transient Expression to Study Gene Expression and Stability 30
Agrobacterium Background 31
A String of Improvements for Agrobacterium 32
Agrobacterium-- Plant Interactions 33
Reducing Agents 33
Agroinfiltration 34
Arabidopsis Floral Dipfloral dip 34
Particle BombardmentParticle bombardment 35
Gene Guns 36
Optimization of DNA Delivery 36
Control of DNA Integration Patternintegration patterns 37
Other Direct DNA Uptakedirect DNA uptake Approaches 37
Protoplasts 38
Whole Tissue Electroporation 38
Silicon Carbide Whiskers 39
Nanofiber ArraysNanofiber arrays 39
Pollen Tube PathwayPollen tube pathway 40
Evidence for Transformation 41
DNA Presence 41
Gene Expression 42
Conclusions 43
References 43
Chapter 2: Plastid Transformation 47
Introduction 47
Delivery of Transforming DNA to the Chloroplast 48
Vector Design 51
Flanking Regions 51
Promoters and UTRs 53
Transgene Stacking and Control of Gene Expression 54
Selection 55
Antibiotic Resistance Markers 55
Other Selection Markers 56
Marker Gene Excision 56
Analysis 57
Conclusions 58
References 58
Chapter 3: Concepts of Marker Genesmarker genes for Plants 62
Introduction 62
Criteria for Choosing the Marker Gene SystemMarker gene system 63
Availability of Selectable Marker GeneSelectable marker gene Systems and Alternatives 65
Positive Selection Marker 65
AntibioticsAntibiotics 65
Neomycin Phosphotransferase IINeomycin phosphotransferase II 66
Hygromycin Phosphotransferase 67
Antibiotic Resistance Genes Beside nptII and hph 67
HerbicidesHerbicides 67
Metabolic Analogous, Toxic, Non-Toxic Agents 68
2-Deoxyglucose-6-Phosphate Phosphatase 68
Xylose Isomerase 69
Isopentenyl Transferaseisopentenyl transferase 69
Phosphomannose Isomerasephosphomannose isomerase 70
Alternative Systems 70
Selectable Marker Gene Eliminationmarker gene elimination 70
Co-TransformationCotransformation 70
Recombinase-Induced EliminationRecombinase induced elimination 72
Transposon-Based EliminationTransposon-based elimination 73
Homologous RecombinationHomologue recombination 74
Screenable Marker GenesScreenable marker genes 74
Negative Selection MarkerNegative selection marker 75
Marker-Free TransformationMarker-free transformation Without Usage of Any Marker Gene 75
Conclusions and Perspective 76
References 77
Chapter 4: Precise Breeding Through All-Native DNA Transformation 84
Introduction 84
Examples of the Intragenic Modification in Potato 85
Requirements for the All-Native DNA Transformation of Potato 88
Intragenic Tomato (S. esculentum): Concentrating the Quality Potential of Tomatotomato into its Fruit 90
Exploring the Diversity of Solanaceous Crops 91
Intragenic Modification of Alfalfa: Optimization of a Forage Feed 92
Exploiting Native Genetic Elements for Canola Oilseed Improvements 93
Drought-Tolerant Perennial Ryegrass 94
Bruise-Tolerant Appleapple 95
Native Markers for Intragenic Transformation 95
Intragenic Crops Are at Least as Safe as Those Developed Through Traditional Methods 96
Conclusions 97
References 97
Chapter 5: Gene Silencing in Plants: Transgenes as Targets and Effectors 101
Introduction 101
Mechanisms of Gene Silencing 102
The Role of Small RNAs 103
MicroRNA Genes 104
Natural Sense--Antisense Gene Pairs 104
Viral Genomic RNA, Transcripts, and Replication Intermediates 104
Heterochromatic and Repetitive Sequences 105
Epigenetic Silencing of Transcription 105
Silencing of Transgene Expression 107
Cis- and Trans-Silencing Of Multi-Copy Transgenes 107
Silencing of Single-Copy Transgenes 110
Reducing the Risk of Transgene Silencing 111
Applications of RNA Interference in Transgenic Plants 112
Applications of RNAi for Crop Protectioncrop protection 114
Applications of RNAi for Crop Improvementcrop improvement and Metabolic Engineering 115
Conclusions 116
References 116
Chapter 6: Breeding with Genetically Modified Plants 124
Genetic Variation in Plant Breeding 124
Breeding Aims 124
Methods for Introducing Transgenes into Elite Plant Material 126
Breeding Methods 128
Line VarietiesLine varieties 129
Open-Pollinated VarietiesOpen pollinated varieties 130
Hybrid Varieties 130
Clone VarietiesClone varieties 132
Safety and Legal Aspects of GMO Breeding 133
Separating Transgenic and Non-Transgenic Breeding Programs 133
Breeding Marker-Free Cultivarsmarker-free cultivars 134
`Cisgenic´ and Transgenic Plants 134
Non-Transgenic Versus Transgenic Breeding 135
Conclusions 136
References 137
Chapter 7: Detection of Genetically Modified Plants in Seeds, Food and Feed 138
Introduction 138
Techniques Used to Detect a Transgenic Plant 139
DNA-Based DetectionDNA-based detection 139
Polymerase Chain Reaction 140
Conventional Qualitative PCR 140
Quantitative Real-Time PCRQuantitative real-time PCR 141
Alternative DNA-Based Techniques 142
Protein-Based DetectionProtein-based detection 143
Lateral Flow Strip 143
Enzyme-Linked Immunosorbent AssayEnzyme-Linked Immunosorbent Assay (ELISA) 144
Method Validationvalidation and Standardisation 144
Detection Strategies 145
ScreeningScreening 145
IdentificationIdentification 149
Quantification 150
Detection of Stacked Events 150
Detection of Unauthorised/Unknown GMOs 151
Method Databases 152
SamplingSampling Issues 152
Conclusions 153
References 153
Part B: Selected Characters of Transgenic Plants and Their Application in Plant Production 158
Chapter 8: Drought Stress Tolerance 159
Introduction 159
Transgenic Plant Strategies for Enhanced Drought Stress Tolerance in Crop Plants 160
OsmoprotectantsOsmoprotectants and Metabolite Engineeringmetabolite engineering 161
Amino Acid-Derived Osmoprotectants 165
ProlineProline 165
Glycine BetaineGlycine betaine 165
Sugar-Related Osmoprotectants 166
FructansFructans 166
Polyols 166
TrehaloseTrehalose 166
Regulatory and Signalling Genessignalling genes: Tools to Engineer Drought Stress Tolerance 167
DREB/CBF: a Landmark Discovery in the Manipulation of Abiotic Stress Tolerance 167
SNAC1/2: Stress-Responsive Plant-Specific Transcription Factors with Distinct Mechanisms of Action 169
HARDY: Engineering Water Use Efficient Rice 170
HD-START: a Developmental Regulator Conferring Drought Tolerance 171
Plant Nuclear Factor Y B Subunits: Field-Validated Drought Tolerance in Maize 171
Rice Calcium-Dependent Protein Kinase 7: Multiple Abiotic Stress Tolerance with Minimal Pleiotropic Events 172
Tobacco Protein Kinase: Sustained Yield in Maize under Water-Limited Conditions 172
Future Prospects: ``Climate-Ready´´ Crops 173
References 174
Chapter 9: Herbicide Resistance 178
Introduction 178
Overview of Adoption 179
Types of Herbicide Resistance 179
Modes of Herbicide Action in Herbicide-Resistant Crops 180
Implications of Genetically Modified Herbicide-Resistant Crops 181
Selection Pressure Indirectly Attributable to Genetically Modified Herbicide-Resistant Crops 181
Evolved Herbicide Resistance 182
Changes in Herbicide Use 182
Lack of Integrated Weed Management 183
Specific Crops with Herbicide Resistance 183
Maize 183
Soybean 184
Cotton 184
Canola 184
Sugarbeets 185
Turf 185
Alfalfa 185
Rice 185
Wheat 186
Implications of Genetically Modified Herbicide Resistance on Cropping Systems 186
Tillage 186
Diversity of Weed Management Tactics 187
Timelines of Weed Management Tactics 188
Herbicide-Resistant Weeds 188
Weedy Near-Relatives to Genetically Modified Herbicide-Resistant Crops -- Gene Flow 189
Implications of Herbicide Resistance -- Persistence in the Agroecosystem 190
Conclusions 191
References 192
Chapter 10: Insectinsect and Nematodenematode Resistance 196
Introduction 196
R Gene-Mediated Resistance 197
Plant ResistancePlant resistance and Resistance Gene 197
Plant Parasite Resistance and Resistance Genes 198
Significance and Limitations of Plant Resistance Genes 200
Engineering of Insect and Nematode Resistance 201
Anti-Insect/Nematode Genes 202
Bt Toxins 202
Proteinase InhibitorsProteinase Inhibitors 203
LectinsLectins 205
a-Amylase Inhibitors 206
ChitinasesChitinases and Others 206
RNA Interference-Based Gene Silencing 207
Conclusions 208
References 208
Chapter 11: Metabolic Engineering 217
Introduction 217
Strategies for Metabolic Engineering in Plants 218
Engineering of Primary Metabolism 219
Carbohydrate Metabolism 219
Production of Novel Carbohydrates in Transgenic Plants 222
Metabolic Engineering of Lipid Metabolismlipid metabolism 224
Biosynthesis of Storage Lipidsstorage lipids 224
Genetic Engineering of Plant Lipid Metabolism 226
Unusual Medium-Chain Fatty Acids 226
Unusual Long-Chain Fatty AcidsUnusual long-chain fatty acids 227
Fatty Acids with Additional Functional Groups 229
Engineering of Secondary Metabolism for Human Health and Nutrition 230
Flavonoids 230
Vitamins 231
Conclusions 232
References 232
Chapter 12: Pharmaceuticals 238
Introduction 238
Expression Systems 239
Transient Expression Systems 239
Stable Expression Systems 240
Transplastomic Plants 240
Nuclear Transgenic Plants 242
Post-Translational Modifications 243
Downstream Processing 245
PMPs in Advanced Development 245
Glucocerebrosidase 245
Insulin 246
Idiotype Vaccines 247
Interferon 248
Conclusion 248
References 249
Chapter 13: Biopolymers 253
Introduction 253
Transgene-Encoded Biopolymers 254
Starch and Cellulose 255
Starch 255
Cellulose 255
Glucoside 257
Polyhydroxyalkanoates 258
Protein-Based Biomaterials 259
Fibrous Proteins 259
Non-Ribosomally Produced Poly-Amino Acids 261
Conclusion 263
References 264
Chapter 14: Engineered Male Sterility 269
Introduction 269
Natural Male Sterility Systems in Plants 270
Cytoplasmic Male Sterility 270
Nuclear Male Sterility 271
Methods of Producing Male-Sterile Plants 272
The Selective Destruction of Tissues Important for the Production of Functional Pollenpollen 272
Changing the Levels of Metabolites Needed for the Production of Viable Pollenpollen 274
Engineering Cytoplasmic Male-Sterile Plants 275
Strategies for the Multiplication of Male-Sterile Lines 275
Herbicide Application for Selection of Male-Sterile Plants 276
Reversible Male Sterility 276
Use of Maintainermaintainer Lines 277
Commercial Use of Male Sterility 277
Conclusions and Future Perspectives 277
References 278
Part C: Transgenic Plants in Breeding and Crop Production 282
Chapter 16: Triticeae Cereals 300
Introduction 300
The Generation of Transgenic Triticeae Plants 301
Transgene Expression Systems 302
Tolerance to Abiotic Stressabiotic stress 303
DroughtDrought and Salinitysalinity 304
Aluminium Toxicity 305
Resistance to Fungal Infection 305
Regulators of Plant Defence 306
Pathogenesis-Related Proteins 306
R Proteins 308
Fungal Proteins 309
Viral Proteins 309
Resistance to Viral Infection 309
Resistance to Insects 310
Grain Quality 310
Production of Recombinant Proteins 312
References 313
Chapter 17: Fruit CropsFruit Crops 320
Introduction 320
Temperate Fruit CropsTemperate fruit crops 321
Top FruitTop fruit 321
MalusMalus Species (AppleApple) 321
PyrusPyrus Species (PearPear) 327
PrunusPrunus Species (AlmondAlmond, Apricotapricot, Sweet and Sour Cherrycherry, Cherry Rootstockscherry rootstocks, Peachpeac 329
Small FruitSmall fruit 332
FragariaFragaria Species (StrawberryStrawberry) 332
GrapevineGrapevine 333
RibesRibes Species (Blackcurrantblackcurrant, Redcurrantredcurrant, Gooseberrygooseberry) 336
RubusRubus Species (RaspberryRaspberry, Blackberryblackberry) 336
VacciniumVaccinium Species (BlueberryBlueberry, Cranberrycranberry) 336
Tropical and Subtropical Fruit CropsTropical and subtropical fruit crops 337
AvocadoAvocado 337
BananaBanana 337
CitrusCitrus Species 338
KiwifruitKiwifruit 340
MangoMango 340
PapayaPapaya 341
PersimmonPersimmon 342
PineapplePineapple 342
References 343
Chapter 18: Maize 362
Introduction 362
Culture MediaCulture Media and Supplements 363
GenotypeGenotype 364
Explant 364
Transformation 365
Free DNA DeliveryFree DNA Delivery in Protoplasts 365
Intact Tissue Electroporation 366
Silicon Carbide 366
Microprojectile BombardmentMicroprojectile Bombardment 367
AgrobacteriumAgrobacterium 371
BenefitsBenefits 374
References 376
Chapter 19: Ornamentals 381
Introduction 381
Flower Colour ModificationsFlower colour modifications 382
Red and Pink Flowers 384
Yellow and Orange Flowers 384
Blue Flowers 385
White Flowers 385
Pigmentation PatternsPigmentation patterns 385
Postharvest QualityPostharvest quality 386
Plant ArchitecturePlant architecture 388
Disease ResistanceDisease resistance 390
Virus ResistanceVirus resistance 391
Resistance Against Fungi and BacteriaResistance against fungi and bacteria 392
Insect ResistanceInsect resistance 393
Flowering TimeFlowering time 394
Modification of Flower Structureflower structure 394
Improvement of Abiotic Stress Toleranceabiotic stress tolerance 395
Modification of Floral Scentfloral scent 396
Conclusion 397
References 397
Chapter 20: Potato 404
Introduction 404
Pathogen Resistance 405
InsectsInsect resistance 405
VirusesVirus resistance 406
Phytophthora infestansPhytophthora infestans 407
Tuber Quality Traits 408
Blackspot Bruise 408
Cold-Induced SweeteningCold-induced sweetening 408
Nutritional Value 409
Amino Acids/Protein 409
Carotenoids 410
Fructan/Inulin 411
Production of Biopolymers 412
Starch 412
Polyhydroxyalkanoates 412
Cyanophycin/Poly-Aspartate 413
Spider Silk 414
Conclusions 414
References 415
Chapter 21: Rapeseed/Canola 420
Introduction 420
Transformation Using Direct Gene Transfer Methods 421
Transformation Using Agrobacterium tumefaciens 421
Explant Type, Additives and Genotype Dependance 421
Agrobacterium Strains 422
Transformation Using Protoplasts 423
Transformation Using Haploids 423
Transformation Avoiding Tissue Culture 424
Plant-Selectable Marker Genes and Marker Gene-Free Transgenic Plants 424
Employment of Transgenic Oilseed Rape in Breeding 425
Employment of Transgenic Oilseed Rape in Crop Production 428
Conclusions 430
References 430
Chapter 22: Rice 434
Introduction 434
Rice Transformation Technology and Functional GenomicsFunctional genomics 435
Insecticidal Rice 436
Bt Rice 436
GNA Rice 437
Disease-Resistant Rice 438
Resistance to Bacterial Blight 438
Resistance to Fungal Diseases 438
Resistance to Viral Diseases 439
Abiotic Stress ToleranceAbiotic stress tolerance 440
Quality ImprovementQuality improvement 444
Nutrient-Use Efficiency 445
Nitrogen-Use EfficiencyNitrogen-use efficiency 445
Phosphorus-Use EfficiencyPhosphorus-use efficiency 446
Yield 448
Herbicide-Tolerant Rice 450
Prospects 451
References 452
Chapter 23: Sugarcane 463
Introduction 463
Origin 463
Sugarcane Breeding, Biotechnology and Biosafety 464
In Vitro Culture 465
In Vitro Culture for Sugarcane Improvement 465
Sugarcane Somatic Embryogenesis 466
Sugarcane Organogenesis 466
Genetic Engineering of Sugarcane 467
Methods of Transformation 467
Selection of the Transformed Tissues 471
Traits of Interest 471
Herbicide Resistance 471
Biotic Stress Tolerance 472
Abiotic Stress Tolerance 474
Metabolic Engineering of the Carbohydrate Metabolism and Value-Added Products 475
Future Trends 476
References 477
Chapter 24: Soybean 483
Introduction 483
Methodology 484
Cot Node and other Organogenic Transformation Systems 484
Cot Node 484
Stem Node 484
Bombardment of the Shoot Tip 485
Embryogenic Culture Transformation System 485
Whole-Plant Transformation Systems 486
Floral Dip 486
Pollen Tube Pathway 486
Composite Plants 486
Virus-Induced Gene Silencing 487
Other Considerations for Transformation 487
Multi-Gene InsertionsMulti-gene insertions and Marker-Free PlantsMarker-free plants 487
Selectable Markers 488
Homozygosity DeterminationHomozygosity determination 489
Applications of Transformation Technology 490
Herbicide ResistanceHerbicide Resistance 490
Modification of Oil CompositionOil Composition 490
Nematode ResistanceNematode Resistance 492
Isoflavones 493
Insect ResistanceInsect Resistance 493
Disease Resistance 494
PhytasePhytase 494
Seed Protein Compositionprotein composition 495
Gene Discovery and Promoters 496
Genomic Resources for Selection of Promoters and Genes for Modification 496
Promoter EvaluationPromoter Evaluation 497
Characterization of Soybean Promoters 499
Future of Soybean Transformation 500
References 501
Chapter 25: Vegetables 509
Introduction 509
Economically Important Vegetable Families 525
SolanaceaeSolanaceae 525
Solanum lycopersicon L.SolanaceaeSolanum lycopersicon L. 525
Solanum melongena L.SolanaceaeSolanum melongena L. 526
Capsicum annuum L.SolanaceaeCapsicum annuum L. 528
BrassicaceaeBrassicaceae (Brassica oleracea L.BrassicaceaeBrassica oleracea L., B. rapa L.BrassicaceaeBrassica rapa L., Raphan 529
FabaceaeFabaceae (Pisum sativum L.FabaceaePisum sativum L., Phaseolus vulgaris L.FabaceaePhaseolus vulgaris L.) 531
CucurbitaceaeCucurbitaceae [Cucumis sativus L.CucurbitaceaeCucumis sativus L., C. melo L.CucurbitaceaeCucumis melo L., Cucurbi 533
AsteraceaeAsteraceae 534
Lactuca sativa L.AsteraceaeLactuca sativa L. 534
CichoriumAsteraceaeCichorium ssp. intybus L., C. spinosum L., C. endivia L. 535
ApiaceaeApiaceae (Daucus carota L.)ApiaceaeDaucus carota L. 536
Chenopodiaceae (Spinacia oleracea L.Spinacia oleracea L.) 537
LiliaceaeLiliaceae 537
AlliumLiliaceaeAllium ssp. cepa L., A. porrum L., A. sativum L. 537
Asparagus officinalis L.LiliaceaeAsparagus officinalis L. 538
Conclusions 538
References 539
Part D: Risk Assessment and Economic Applications 561
Chapter 26: Regulatory Oversight and Safety Assessment of Plants with Novel Traits 562
Introduction - From Foragers to Genetic Modification in a Genomic Era 562
Regulatory Oversight of GM Plants and Their Derived Food and Feed Products 564
Process-Based Versus Product-Based Approach 564
Regulatory Framework for GMOs in the EU 565
Risk Assessment Principles 566
Interplay of Risk Assessment, Risk Management and Risk Communication 566
Risk Assessment Methodology and Terminology 567
Problem Formulation 568
Assessment Endpoints 568
Conceptual Model 569
Analysis Plan 570
Risk Assessment Principles and Concepts 571
Comparative Risk Assessment and Familiarity Concept 571
Case-by-Case Principle 572
Iterative and Adaptive 572
Tiered Approach 573
EFSA GMO Panel Guidance and Further Prospectives 574
EFSA Scientific Colloquium on Environmental Risk Assessment of GM Plants 575
Self-Tasking Working Group on NTO Testing 576
Update of Environment Sections of the EFSA Guidance on the Risk Assessment of GM Plants and Derived Food and Feed Products 576
Discussion and Conclusions 577
References 580
Chapter 27: Environmental Impact of Genetically Modified Maize Expressing Cry1 Proteins 584
Introduction 584
Potential Unintended Effects on Plant Fitness Due to the Genetic Modification 585
Potential for Gene Transfer 586
Plant to Micro-Organism Gene Transfergene transfer 586
Plant to Plant Gene Transfer 586
Potential Interactions of the GM Plant with Target Organisms 587
Potential Interactions of the GM Plant with Non-Target Organisms 590
Persistence of Cry1 Proteins in Soil: ExposureAssessment 590
Biological Effects in Soil: General Impact Assessment 592
Assessment of Impact on Earthworms 593
Assessment of Impact on Isopods 594
Assessment of Impact on Nematodes 595
Assessment of Impact on Collembolans 596
Cry1 Genes in Water: Exposure Assessment in Aquatic Environments 597
Presence of Cry1 Proteins in Water: Impact Assessment in Aquatic Environmentsaquatic environments 598
Exposure and Impacts on Non-Target LepidopteraLepidoptera 599
Global Analysis of Impacts on Non-TargetEntomofauna 602
Trophic Chain Effects on Predators 603
Trophic Chain Effects on Parasitoids 605
Assessment of Impacts on Pollinating Insects 606
Potential Impacts on Human and Animal Health 608
Potential Interaction with the Abiotic Environment and Biogeochemical Cycles 608
Impacts of the Specific Cultivation, Management and Harvesting Techniques 610
Monitoring 611
Conclusions 612
References 613
Chapter 28: Benefits of Transgenic Plants: A Socioeconomic Perspective 624
Introduction 624
Impacts of Insect-Resistant Crops 625
Agronomic Effects 625
Economic Effects 627
Povertypoverty effects and Distribution Effects 628
Environmental and Health Effects 628
Impacts of Herbicide-Tolerant Crops 631
Agronomic and Economic Effects 631
Environmental Effects 633
Potential Impacts of Future Transgenic Crops 633
Crops with Improved Agronomic Traits 633
Crops with Improved Nutritional Traits 634
Conclusions 635
References 636
Chapter 29: Risk Assessment and Economic Applications - the Cartagena Protocol on BiosafetyCartagena Protocol on Biosafety: GMO 639
29.1Introduction 639
29.2The Cartagena Protocol on Biological Safety 640
29.2.1The Convention on Biological Diversity as the Basis for the Cartagena Protocol on Biological Safety 640
29.2.2The Cartagena Protocol on Biosafety and the Biosafety Clearing HouseBiosafety Clearing House 641
29.3GMO ApprovalGMO approval 645
29.3.1European Union 645
29.3.1.1Authorisation of Genetically Modified Food and Feed in the EU 648
29.3.1.2Placing on the MarketPlacing on the Market of Genetically Modified Organisms According to Directive 2001/18/EC 651
29.3.2United States of America 652
29.4GMO Approval, GMO Labelling and GMO Trade 653
29.5Conclusions 654
References 654
Chapter 30: Public Perceptions of Modern Biotechnology and the Necessity to Improve Communication 657
Introduction 657
Societal Debate and Its Problems 658
Statistic Data on Public Attitudesattitudes Towards Biotechnology 659
Frames of Reference by Promotors and Critics 661
Risk Perception and the Role of Confidenceconfidence and Trusttrust 661
Scepticism Against Technology and Progress? 663
Insufficient Approaches 664
Improvements of Communicationcommunication with the Public 665
Respect to Sustainabilitysustainability and Ethics 667
InvolvementInvolvement of Consumersconsumers 667
References 669
: Index 670