Contents

Cover

Title page

Copyright page

Introduction

Part 1: Genomics

Chapter 1: Exploring Genomics Research in the Context of Some Underutilized Legumes—A Review
1. 1.1 Introduction
2. 1.2 Velvet Bean [Mucuna pruriens (L.) DC. var. utilis (Wall. ex Wight)] Baker ex Burck
3. 1.3 Psophocarpus tetragonolobus (L.) DC.
4. 1.4 Vigna umbellata (Thunb.) Ohwiet. Ohashi
5. 1.5 Lablab purpureus (L.) Sweet
6. 1.6 Avenues for Future Research
7. 1.7 Conclusions
8. Acknowledgments
9. References
Chapter 2: Overview of Insecticidal Genes Used in Crop Improvement Program
1. 2.1 Introduction
2. 2.2 Insect-Resistant Transgenic Model Plant
3. 2.3 Insect-Resistant Transgenic Dicot Plants
4. 2.4 Insect-Resistant Transgenic Monocot Plants
5. 2.5 Working Principle of Insecticidal Genes Used in Transgenic Plant Preparation
6. 2.6 Discussion
7. References
Chapter 3: Advances in Crop Improvement: Use of miRNA Technologies for Crop Improvement
1. 3.1 Introduction
2. 3.2 Discovery of miRNAs
3. 3.3 Evolution and Organization of Plant miRNAs
4. 3.4 Identification of Plant miRNAs
5. 3.5 miRNA vs. siRNA
6. 3.6 Biogenesis of miRNAs and Their Regulatory Action in Plants
7. 3.7 Application of miRNA for Crop Improvement
8. 3.8 Concluding Remarks
9. References
Chapter 4: Gene Discovery by Forward Genetic Approach in the Era of High-Throughput Sequencing
1. 4.1 Introduction
2. 4.2 Mutagens Differ for Type and Density of Induced Mutations
3. 4.3 High-Throughput Sequencing is Getting Better and Cheaper
4. 4.4 Mapping-by-Sequencing
5. 4.5 Different Mapping Populations for Specific Need
6. 4.6 Effect of Mutagen Type on Mapping
7. 4.7 Effect of Bulk Size and Sequencing Coverage on Mapping
8. 4.8 Challenges in Variant Calling
9. 4.9 Cases Where Genome Sequence is either Unavailable or Highly Diverged
10. 4.10 Bioinformatics Tools for Mapping-by-Sequencing Analysis
11. Acknowledgments
12. References
Chapter 5: Functional Genomics of Thermotolerant Plants
1. 5.1 Introduction
2. 5.2 Functional Genomics in Plants
3. 5.3 Thermotolerant Plants
4. 5.4 Studies on Functional Genomics of Thermotolerant Plants
5. 5.5 Concluding Remarks
6. Abbreviations
7. References

Part 2: Metabolomics

Chapter 6: A Workflow in Single Cell-Type Metabolomics: From Data Pre-Processing and Statistical Analysis to Biological Insights
1. 6.1 Introduction
2. 6.2 Methods and Data
3. 6.3 Results
4. 6.4 Discussion
5. 6.5 Conclusion
6. Conflicts of Interest
7. Acknowledgment
8. References
Chapter 7: Metabolite Profiling and Metabolomics of Plant Systems Using ¹H NMR and GC-MS
1. 7.1 Introduction
2. 7.2 Materials and Methods
3. 7.3 Selected Applications of Metabolomics and Metabolite Profiling
4. Acknowledgments
5. Competing Interests
6. References
Chapter 8: OMICS-Based Approaches for Elucidation of Picrosides Biosynthesis in Picrorhiza kurroa
1. 8.1 Introduction
2. 8.2 Cross-Talk of Picrosides Biosynthesis Among Different Tissues of P. kurroa
3. 8.3 Strategies Used for the Elucidation of Picrosides Biosynthetic Route in P. kurroa
4. 8.4 Strategies Used for Shortlisting Key/Candidate Genes Involved in Picrosides Biosynthesis
5. 8.5 Complete Architecture of Picrosides Biosynthetic Pathway
6. 8.6 Challenges and Future Perspectives
7. Abbreviations
8. References
Chapter 9: Relevance of Poly-Omics in System Biology Studies of Industrial Crops
1. 9.1 Introduction
2. 9.2 System Biology of Crops
3. 9.3 Industrial Crops
4. 9.4 Poly-Omics Application in System Biology Studies of Industrial Crops
5. 9.5 Concluding Remarks
6. Abbreviations
7. References

Part 3: Bioinformatics

Chapter 10: Emerging Advances in Computational Omics Tools for Systems Analysis of Gramineae Family Grass Species and Their Abiotic Stress Responsive Functions
1. 10.1 Introduction
2. 10.2 Gramineae Family Grass Species
3. 10.3 Abiotic Stress
4. 10.4 Emerging Sequencing Technologies
5. 10.5 Omics Resource in Poaceae Species
6. 10.6 Role of Functional Omics in Dissecting the Stress Physiology of Gramineae Members
7. 10.7 Systems Analysis in Gramineae Plant Species
8. 10.8 Nutritional Omics of Gramineae Species
9. 10.9 Future Prospects
10. 10.10 Conclusion
11. Acknowledgments
12. References
Chapter 11: OMIC Technologies in Bioethanol Production: An Indian Context
1. 11.1 Introduction
2. 11.2 Indian Scenario
3. 11.3 Cellulolytic Enzymes Producing Bacterial Strains Isolated from India
4. 11.4 Biomass Sources Native to India
5. 11.5 Omics Data and Its Application to Bioethanol Production
6. 11.6 Conclusion
7. References

Part 4: Advances in Crop Improvement: Emerging technologies

Chapter 12: Genome Editing: New Breeding Technologies in Plants
1. 12.1 Introduction: Genome Editing
2. 12.2 GE: The Basics
3. 12.3 Engineered Nucleases: The Key Players in GE
4. 12.4 Targeted Mutations and Practical Considerations
5. 12.5 New Era: CRISPR/Cas9
6. 12.6 GE for Improving Economic Traits
7. 12.7 Biosafety of GE Plants
8. 12.8 What’s Next: Prospects
9. References
Chapter 13: Regulation of Gene Expression by Global Methylation Pattern in Plants Development
1. 13.1 Introduction
2. 13.2 Nucleic Acid Methylation Targets in the Genome
3. 13.3 Nucleic Acid Methyl Transferase (DNMtase)
4. 13.4 Genomic DNA Methylation and Expression Pattern
5. 13.5 Pattern of DNA Methylation in Early Plant Life
6. 13.6 DNA Methylation Pattern in Mushroom
7. 13.7 Methylation Pattern in Tumor
8. 13.8 DNA Methylation Analysis Approaches
9. References
Chapter 14: High-Throughput Phenotyping: Potential Tool for Genomics
1. 14.1 Introduction
2. 14.2 Relation of Phenotype, Genotype and Environment
3. 14.3 Features of HTP
4. 14.4 HTP Pipeline and Platforms
5. 14.5 Controlled Environment-Based Phenotyping
6. 14.6 Field-Based High-Throughput Plant Phenotyping (Fb-HTPP)
7. 14.7 Applications of HTP
8. 14.8 Conclusion and Future Thrust
9. References

Index

End User License Agreement

List of Illustrations

Chapter 1

Figure 1.1 Variability for seed characters in M. pruriens germplasm....
Figure 1.2 (a) Flowering and (b) fruiting in P. tetragonolobus (L.) DC....

Chapter 3

Figure 3.1 The plant miRNA biogenesis [17]....

Chapter 4

Figure 4.1 Schematic flowchart of mapping by sequencing involving isogenic mapping....
Figure 4.2 Schematic plots of allele frequency of induced mutations along a chromosome....
Figure 4.3 Effect of pool size (or bulk size) and sequencing coverage on the interval size....

Chapter 5

Figure 5.1 Model summarizing the up-stream and down-stream changes governing....

Chapter 6

Figure 6.1 Summarized statistical workflow (normalization, transformation, and scaling....
Figure 6.2 Displaying the metabolic pathway enrichment analysis output for the metabolites....
Figure 6.3 Multivariate analysis displaying the best combinations of normalization....
Figure 6.4 (a) A four-way Venn diagram showing the unique and shared metabolites between....
Figure 6.5 Box whisker plots showing the output of 12 normalization methods for (A) C....
Figure 6.6 MDS plots: (A) control, (B) treatment; where numbers 1–6 represent the six....

Chapter 7

Figure 7.1 A typical experimental workflow showcasing the metabolomics of plants. The....
Figure 7.2 Steps for ¹H NMR data analysis—1. Metabolite extraction: Sample....
Figure 7.3 Steps for GC-MS data analysis—1. Sample preparation by homogenizing....
Figure 7.4 Data interpretation using various statistical approaches—different....
Figure 7.5 Representation of pathway mapping using MetaboAnalyst: the extent of impact....

Chapter 8

Figure 8.1 Bioactivities, sites of biosynthesis/accumulation in P. kurroa....
Figure 8.2 Overview of strategies used for the shortlisting of key picrosides biosynthetic....
Figure 8.3 Complete metabolic architecture of P-I and P-II biosynthesis in P. kurroa....

Chapter 9

Figure 9.1 Omics data in relation with system biology. (Adapted from Ali et al.,....

Chapter 10

Figure 10.1 Illustration of C3 and C4 stress responsive gene families and their associated....
Figure 10.2 A simplified omic space with advanced omics layers....
Figure 10.3 Deciphering the nutritional traits in Gramineae species by integrated omics....

Chapter 11

Figure 11.1 Schematic diagrams explaining the bioethanol production from second-generation....

Chapter 12

Figure 12.1 Major DNA repair pathways in plants. (a) Classical NHEJ; (b) microhomology-based....
Figure 12.2 Types of different engineered nucleases used for GE, namely, ZFN, TALEN....
Figure 12.3 Historical timeline of the development of CRISPR/Cas technique [131]....
Figure 12.4 Intended modifications using GE in plants....
Figure 12.5 Basic strategies and important practical considerations for the GE in plants....

Chapter 13

Figure 13.1 Schematic of packaging of eukaryotic genome and histone modification associated....
Figure 13.2 Methylation of nucleic acid on cytosine nucleotide site....
Figure 13.3 Schematic classification of DNA methyltransferase in plants. DNMtases....
Figure 13.4 Schematic diagram of DNA methylation pattern. Expression of genes....
Figure 13.5 Schematic diagram of ribonucleic acid mediated methylation. SUVH2/9 component....
Figure 13.6 Flowchart for the analysis of global DNA methylation in genomes....
Figure 13.7 Flowchart for bioinformatics analysis of whole genome methylation....

Chapter 14

Figure 14.1 Relationship between genotype, phenotype, and environment....
Figure 14.2 Flowchart for HTP....
Figure 14.3 Applications of HTP....

List of Tables

Chapter 1

Table 1.1 Comparison of genomic resources in four lesser-known legume species....

Chapter 2

Table 2.1 Insect-resistant transgenic model plant....
Table 2.2 Insect-resistant transgenic dicot plant....
Table 2.3 Insect-resistant transgenic monocot plant....

Chapter 3

Table 3.1 Comparison of miRNAs and siRNAs....
Table 3.2 Plant miRNAs and their biological functions....

Chapter 4

Table 4.1 Differences among three mapping populations typically used....

Chapter 5

Table 5.1 RNAi technique application in some agriculturally significant plants....
Table 5.2 List of thermotolerant plants region-wise....

Chapter 6

Table 6.1 Data showing the effect of various normalization, transformation, and scaling....
Table 6.2 Data showing the effect of various normalization, transformation, and scaling....

Chapter 8

Table 8.1 Details of strategies used to shortlist key candidate genes/transcripts....
Table 8.2 Details of strategies used to shortlist key candidate genes/transcripts....

Chapter 9

Table 9.1 List of industrial crops....

Chapter 10

Table 10.1 Gramene species FAOSTAT report....
Table 10.2 Bioinformatic databases and tools for multiple omics approaches....
Table 10.3 Status of NGS technology in Gramineae family food crops....

Chapter 11

Table 11.1 List of microorganism strains and enzymes isolated reported to be indigenous....
Table 11.2 Availability of genome, proteome, and transcriptome data for biomass sourced....

Chapter 12

Table 12.1 Comparative summary of GE techniques....
Table 12.2 Summary of various types of CRISPR plasmids for plants....
Table 12.3 Summary of available Cas9 variants....
Table 12.4 Comparison between Cas9 and Cpf1 proteins....
Table 12.5 Brief differentiation between RNAi and CRISPR/Cas system....
Table 12.6 Successful application of GE in model and important crop plants....
Table 12.7 Probable target genes for GE in rice....

Chapter 14

Table 14.1 Summary of image-based phenotyping....

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Library of Congress Cataloging-in-Publication Data

ISBN 978-1-119-50993-6

Introduction

Climate change challenges could be tackled with the advent of techniques in plant biotechnology, which is a key component to usher sustainable food production and productivity. Plant biotechnology has been started with the culturing of plant cells in various media, which depend on the totipotency of the plant cell. Further, with the advancement of genetic engineering, introducing foreign genes into cell and tissue has become an important tool to develop genetically modified (GM) transgenic crops with enhanced/improved characteristics and traits. In recent studies, the plant biotechnology domain has been tremendously changed/shifted from GM crops and gene manipulation to “OMICS”-based approaches to decipher the underlying mechanisms for abiotic and biotic stress tolerance. Advances in instrumentation and technologies revealed that the genomics, proteomics, metabolomics, methylome (epigenetic regulation), bioinformatics, and phenomics have great potential for identifying and characterizing novel traits in plants to meet environmental challenges. To understand the underlying tolerant mechanisms for climate change conditions, an attempt has been made to conglomerate all interdisciplinary branches under one umbrella to emphasize the essentiality of inter-allied sciences for tackling the problem.

To meet the nation’s food demand, fusion of improved varieties with superior genetics to seed chain at appropriate time intervals is inevitable. The primary objective of developing new varieties and hybrids of various crop species will only be achieved through embracing new strategies/technologies and practical implementation for increased productivity. This book is a reflection of the role played by new OMICS technologies in improving the food and nutritional security. Moreover, OMICS potential to use resources effectively for sustainable production has been illustrated vividly to understand the roles of newer technologies.

Agricultural scientists are striving toward the development of appropriate technologies in the form of improved varieties with higher yielding capacity, a wide range of adaptability and resistance to multiple pests, apt for complex and diverse agro-ecological situations. Continued innovations in the field of plant breeding along with the availability of modern tools and techniques provided dividends, enabling varietal development at a much higher pace. Accordingly, in Chapter 1, legume resources such as velvet bean, winged bean, rice bean and lablab bean, which are rich in protein, have been studied using a genomics approach to facilitate toward molecular breeding and gene discovery programs in the near future. Chapter 2 mainly emphasizes the dissection of insecticidal genes and their application for crop improvement. In addition, this chapter throws light over genetically modified crops in controlling pests such as BT technology, and expression of enzymes like chitinase has been explored; it is concluded that transgenic technology coupled with integrated pest management could alleviate the pest problem and enhance the crop productivity. In Chapter 3, miRNA (noncoding small endogenous regulatory RNAs) technologies for crop improvement have been placed for appraisal of latest developments in the domain. miRNAs mediate gene silencing (fully or nearly complementary targets) either through cleavage of target mRNA or translational repression in plants. In this domain, right from first identification of miRNA genes, let-7 and lin-4 from Caenorhabditis elegans, thousands of miRNAs have been identified in plants, and the current MiRBase entries for plants (viridiplantae) have 10,504 mature sequences, which indicates that these molecules could play a prominent role in crop improvement.

Forward genetics approaches are very appealing than conventional genetics; as a result, few chapters are dedicated to forward genetic approaches and their utility in identifying a gene function, investigating the causal locus/gene, and developing thermotolerant plants (see Chapters 4 and 5). Besides genomics and forward genetics approaches, metabolomics is a new branch, which is emerging and helpful to understand the stress-tolerant mechanisms in the plants. As this field is new, Chapter 6 has been focused on metabolomics methodology using single cell type such as the stomatal guard cells that have been used for analyses of the stress-responsive metabolomes when challenged with stressors. Using bioinformatics and statistical approaches, the dataset of explored guard cell metabolome response to a given treatment (bicarbonate) could decipher the cellular mechanisms pertaining to biological events. In Chapter 7, regular procedures for metabolite profiling and metabolomics analysis in plant systems using proton nuclear magnetic resonance (¹H NMR) spectroscopy and gas chromatography–mass spectroscopy (GC-MS) have been elucidated. Further, explanation on general experimental workflow, metabolite data acquisition, metabolite profiling, and statistical analysis for metabolomics using MetaboAnalyst have been dealt with.

OMICS techniques have been dynamic and possess massive potential for crop improvement, which is yet to be reaped particularly in medicinal plants. Application of OMICS in the identification of alkaloids and other secondary metabolites could aid in developing novel insecticides. Chapters 8, 9, and 11 particularly deal with OMICS and poly-OMICS approaches not only on medicinal plants but also on systems biology and biofuel production. Moreover, bioinformatics amalgamating with recent techniques such as CRISPR-Cas9, methylome (epigenetic regulation), and phenomics could be more promising and have been duly accorded in Chapters 10, 12, 13, and 14. It is noteworthy to observe that genome editing (GE), a new breeding technology (NBT), has shown potential to transform not only in fundamental research of plant biology but more importantly also for addressing growing challenges of food security. Hence, due accord has been given to introduce the basic concepts of genome editing with CRISPR-Cas9 in Chapter 12. Regulation of copy number of genes has been maintained by methylation pattern and is considered as epigenetic regulation. Recent reports reveal that nucleic acid methylation in plants like Arabidopsis, maize, and rice has shown that H3K9me2-dependent pathway, ribonucleic acid directed nucleic acid methylation pathway, and mobile siRNAs are the key pathways in the regulation of gene copy number, which is explained in Chapter 13. In addition to these domains, phenomics field is emerging, which has great potential to determine the physiological changes occurring in the plants in response to metabolites and physical factors, and also helps in the development of microfluidic devices. To appraise the advancements taking place in the field, Chapter 14 deals about the know-how, interpretation, and applications in plant biology and crop improvement.

This book aims to keep abreast with the advances taking place in OMICS studies that ultimately aid in confronting climatic challenges. Unprecedentedly, this book is diverse, encompassing several chapters with the latest information, emphasizing new aspects. Indeed, this book would be helpful to plant biotechnologists, plant breeders, agricultural biotechnologists, policymakers, and plant physiologists. Students could refer to this book for competitive exams. It is hoped that this book will be an enriching reference material.

OMICS-Based Approaches in Plant Biotechnology

Introduction

Part 1
GENOMICS