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Introduction xiii Part 1: Genomics 1 1 Exploring Genomics Research in the Context of Some Underutilized Legumes-A Review 3 Patrush Lepcha, Pittala Ranjith Kumar and N. Sathyanarayana 1.1 Introduction 3 1.2 Velvet Bean [Mucuna pruriens (L.) DC. var. utilis (Wall. ex Wight)] Baker ex Burck 4 1.3 Psophocarpus tetragonolobus (L.) DC. 7 1.4 Vigna umbellata (Thunb.) Ohwiet. Ohashi 8 1.5 Lablab purpureus (L.) Sweet 9 1.6 Avenues for Future Research 10 1.7 Conclusions 12 Acknowledgments 12 References 12 2 Overview of Insecticidal Genes Used in Crop Improvement Program 19 Neeraj Kumar Dubey, Prashant Kumar Singh, Satyendra Kumar Yadav and Kunwar Deelip Singh 2.1 Introduction 19 2.2 Insect-Resistant Transgenic Model Plant 21 2.3 Insect-Resistant Transgenic Dicot Plants 27 2.4 Insect-Resistant Transgenic Monocot Plants 34 2.5 Working Principle of Insecticidal Genes Used in Transgenic Plant Preparation 39 2.6 Discussion 41 References 42 3 Advances in Crop Improvement: Use of miRNA Technologies for Crop Improvement 55 Clarissa Challam, N. Nandhakumar and Hemant Balasaheb Kardile 3.1 Introduction 56 3.2 Discovery of miRNAs 56 3.3 Evolution and Organization of Plant miRNAs 57 3.4 Identification of Plant miRNAs 58 3.5 miRNA vs. siRNA 59 3.6 Biogenesis of miRNAs and Their Regulatory Action in Plants 60 3.7 Application of miRNA for Crop Improvement 61 3.8 Concluding Remarks 62 References 70 4 Gene Discovery by Forward Genetic Approach in the Era of High-Throughput Sequencing 75 Vivek Thakur and Samart Wanchana 4.1 Introduction 75 4.2 Mutagens Differ for Type and Density of Induced Mutations 76 4.3 High-Throughput Sequencing is Getting Better and Cheaper 77 4.4 Mapping-by-Sequencing 77 4.5 Different Mapping Populations for Specific Need 81 4.6 Effect of Mutagen Type on Mapping 83 4.7 Effect of Bulk Size and Sequencing Coverage on Mapping 83 4.8 Challenges in Variant Calling 85 4.9 Cases Where Genome Sequence is either Unavailable or Highly Diverged 85 4.10 Bioinformatics Tools for Mapping-by-Sequencing Analysis 86 Acknowledgments 87 References 87 5 Functional Genomics of Thermotolerant Plants 91 Nagendra Nath Das 5.1 Introduction 91 5.2 Functional Genomics in Plants 93 5.3 Thermotolerant Plants 94 5.4 Studies on Functional Genomics of Thermotolerant Plants 98 5.5 Concluding Remarks 99 Abbreviations 100 References 100 Part 2: Metabolomics 105 6 A Workflow in Single Cell-Type Metabolomics: From Data Pre-Processing and Statistical Analysis to Biological Insights 107 Biswapriya B. Misra 6.1 Introduction 108 6.2 Methods and Data 109 6.2.1 Source of Data 109 6.2.2 Processing of Raw Mass Spectrometry Data 109 6.2.3 Statistical Analyses 109 6.2.4 Pathway Enrichment and Clustering Analysis 110 6.3 Results 110 6.3.1 Design of the Study and Data Analysis 110 6.3.2 The Guard Cell Metabolomics Dataset 110 6.3.3 Multivariate Analysis for Insights into Data Pre-Processing 113 6.3.4 Effect of Data Normalization Methods 119 6.4 Discussion 122 6.5 Conclusion 124 Conflicts of Interest 124 Acknowledgment 125 References 125 7 Metabolite Profiling and Metabolomics of Plant Systems Using 1H NMR and GC-MS 129 Manu Shree, Maneesh Lingwan and Shyam K. Masakapalli 7.1 Introduction 129 7.2 Materials and Methods 131 7.2.1 1H NMR-Based Metabolite Profiling of Plant Samples 132 7.2.1.1 Metabolite Extraction 132 7.2.1.2 1H NMR Spectroscopy 132 7.2.1.3 Qualitative and Quantitative Analysis of NMR Signals 134 7.2.2 Gas Chromatography-Mass Spectroscopy (GC-MS) Based Metabolite Profiling 134 7.2.2.1 Sample Preparation 134 7.2.2.2 GC-MS Data Acquisition 135 7.2.2.3 GC-MS Data Pretreatment and Metabolite Profiling 136 7.2.2.4 Validation of Identified Metabolites 136 7.2.3 Multivariate Data Analysis 137 7.3 Selected Applications of Metabolomics and Metabolite Profiling 139 Acknowledgments 140 Competing Interests 140 References 140 8 OMICS-Based Approaches for Elucidation of Picrosides Biosynthesis in Picrorhiza kurroa 145 Varun Kumar 8.1 Introduction 146 8.2 Cross-Talk of Picrosides Biosynthesis Among Different Tissues of P. kurroa 148 8.3 Strategies Used for the Elucidation of Picrosides Biosynthetic Route in P. kurroa 148 8.3.1 Retro-Biosynthetic Approach 149 8.3.2 In Vitro Feeding of Different Precursors and Inhibitors 149 8.3.3 Metabolomics of Natural Variant Chemotypes of P. kurroa 150 8.4 Strategies Used for Shortlisting Key/Candidate Genes Involved in Picrosides Biosynthesis 151 8.4.1 Comparative Genomics 151 8.4.2 Differential Next-Generation Sequencing (NGS) Transcriptomes and Expression Levels of Pathway Genes Vis-a-Vis Picrosides Content 152 8.5 Complete Architecture of Picrosides Biosynthetic Pathway 153 8.6 Challenges and Future Perspectives 161 Abbreviations 162 References 163 9 Relevance of Poly-Omics in System Biology Studies of Industrial Crops 167 Nagendra Nath Das 9.1 Introduction 167 9.2 System Biology of Crops 169 9.3 Industrial Crops 171 9.4 Poly-Omics Application in System Biology Studies of Industrial Crops 176 9.5 Concluding Remarks 177 Abbreviations 177 References 178 Part 3: Bioinformatics 183 10 Emerging Advances in Computational Omics Tools for Systems Analysis of Gramineae Family Grass Species and Their Abiotic Stress Responsive Functions 185 Pandiyan Muthuramalingam, Rajendran Jeyasri, Dhamodharan Kalaiyarasi, Subramani Pandian, Subramanian Radhesh Krishnan, Lakkakula Satish, Shunmugiah Karutha Pandian and Manikandan Ramesh 10.1 Introduction 186 10.2 Gramineae Family Grass Species 187 10.2.1 Oryza sativa 187 10.2.2 Setaria italica 187 10.2.3 Sorghum bicolor 188 10.2.4 Zea mays 188 10.3 Abiotic Stress 188 10.4 Emerging Sequencing Technologies 198 10.4.1 NGS-Based Genomic and RNA Sequencing 199 10.4.2 Tanscriptome Analysis Based on NGS 200 10.4.3 High-Throughput Omics Layers 201 10.5 Omics Resource in Poaceae Species 202 10.6 Role of Functional Omics in Dissecting the Stress Physiology of Gramineae Members 203 10.7 Systems Analysis in Gramineae Plant Species 204 10.8 Nutritional Omics of Gramineae Species 205 10.9 Future Prospects 205 10.10 Conclusion 206 Acknowledgments 207 References 207 11 OMIC Technologies in Bioethanol Production: An Indian Context 217 Pulkit A. Srivastava and Ragothaman M. Yennamalli 11.1 Introduction 217 11.2 Indian Scenario 219 11.3 Cellulolytic Enzymes Producing Bacterial Strains Isolated from India 220 11.3.1 Bacillus Genus of Lignocellulolytic Degrading Enzymes 222 11.3.2 Bhargavaea cecembensis 222 11.3.3 Streptomyces Genus for Hydrolytic Enzymes 230 11.4 Biomass Sources Native to India 230 11.4.1 Albizia lucida (Moj) 230 11.4.2 Areca catechu (Betel Nut) 231 11.4.3 Arundo donax (Giant Reed) 231 11.4.4 Pennisetum purpureum (Napier Grass) 231 11.4.5 Brassica Family of Biomass Crops 231 11.4.6 Cajanus cajan (Pigeon Pea)/Cenchrus americanus (Pearl Millet)/Corchorus capsularis (Jute)/ Lens culinaris (Lentil)/Saccharum officinarum (Sugarcane)/Triticum sp. (Wheat)/Zea mays (Maize) 232 11.4.7 Medicago sativa (Alfalfa) 232 11.4.8 Manihot esculenta (Cassava)/Salix viminalis (Basket Willow)/Setaria italica (Foxtail Millet)/ Setaria viridis (Green Foxtail) 232 11.4.9 Vetiveria zizanioides (Vetiver or Khas) 232 11.4.10 Millets and Sorghum bicolor (Sorghum) 233 11.5 Omics Data and Its Application to Bioethanol Production 233 11.6 Conclusion 239 References 239 Part 4: Advances in Crop Improvement: Emerging Technologies 245 12 Genome Editing: New Breeding Technologies in Plants 247 Kalyani M. Barbadikar, Supriya B. Aglawe, Satendra K. Mangrauthia, M. Sheshu Madhav and S.P. Jeevan Kumar 12.1 Introduction: Genome Editing 248 12.2 GE: The Basics 249 12.2.1 Nonhomologous End-Joining (NHEJ) 250 12.2.2 Homology Directed Repair (HR) 251 12.3 Engineered Nucleases: The Key Players in GE 251 12.3.1 Meganucleases 251 12.3.2 Zinc-Finger Nucleases 256 12.3.3 Transcription Activator-Like Effector Nucleases 257 12.3.4 CRISPR/Cas System: The Forerunner 258 12.4 Targeted Mutations and Practical Considerations 259 12.4.1 Targeted Mutations 259 12.4.2 Steps Involved 260 12.4.2.1 Selection of Target Sequence 261 12.4.2.2 Designing Nucleases 262 12.4.2.3 Transformation 263 12.4.2.4 Screening for Mutation 264 12.5 New Era: CRISPR/Cas9 264 12.5.1 Vector Construction 264 12.5.2 Delivery Methods 266 12.5.3 CRISPR/Cas Variants 266 12.5.3.1 SpCas9 Nickases (nSpCas9) 266 12.5.3.2 Cas9 Variant without Endonuclease Activity 266 12.5.3.3 FokI Fused Catalytically Inactive Cas9 267 12.5.3.4 Naturally Available and Engineered Cas9 Variants with Altered PAM 268 12.5.3.5 Cas9 Variants for Increased On-Target Effect 268 12.5.3.6 CRISPR/Cpf1 268 12.6 GE for Improving Economic Traits 269 12.6.1 Development of Next-Generation Smart Climate Resilient Crops 271 12.6.2 Breaking Yield Incompatibility Barriers and Hybrid Breeding 271 12.6.3 Creating New Variation through Engineered QTLs 271 12.6.4 Transcriptional Regulation 272 12.6.5 GE for Noncoding RNA, microRNA 272 12.6.6 Epigenetic Modifications 273 12.6.7 Gene Dosage Effect 273 12.7 Biosafety of GE Plants 273 12.8 What’s Next: Prospects 276 References 276 13 Regulation of Gene Expression by Global Methylation Pattern in Plants Development 287 Vrijesh Kumar Yadav, Krishan Mohan Rai, Nishant Kumar and Vikash Kumar Yadav 13.1 Introduction 288 13.2 Nucleic Acid Methylation Targets in the Genome 289 13.3 Nucleic Acid Methyl Transferase (DNMtase) 290 13.4 Genomic DNA Methylation and Expression Pattern 291 13.5 Pattern of DNA Methylation in Early Plant Life 292 13.6 DNA Methylation Pattern in Mushroom 293 13.7 Methylation Pattern in Tumor 294 13.8 DNA Methylation Analysis Approaches 294 13.8.1 Locus-Specific DNA Methylation 295 13.8.2 Genome-Wide and Global DNA Methylation 295 13.8.3 Whole Genome Sequence Analysis by Bioinformatics Analysis 296 References 297 14 High-Throughput Phenotyping: Potential Tool for Genomics 303 Kalyani M. Barbadikar, Divya Balakrishnan, C. Gireesh, Hemant Kardile, Tejas C. Bosamia and Ankita Mishra 14.1 Introduction 304 14.2 Relation of Phenotype, Genotype, and Environment 304 14.3 Features of HTP 306 14.4 HTP Pipeline and Platforms 310 14.5 Controlled Environment-Based Phenotyping 311 14.6 Field-Based High-Throughput Plant Phenotyping (Fb-HTPP) 311 14.7 Applications of HTP 313 14.7.1 Marker-Assisted Selection and QTL Detection 314 14.7.2 Forward and Reverse Genetics 315 14.7.3 New Breeding Techniques 315 14.7.3.1 Envirotyping 315 14.8 Conclusion and Future Thrust 316 References 316 Index 323

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