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Satbir Singh Gosal · Shabir Hussain Wani Editors Biotechnologies of Crop Improvement, Volume 2 Transgenic Approaches Biotechnologies of Crop Improvement, Volume 2 Satbir Singh Gosal • Shabir Hussain Wani Editors Biotechnologies of Crop Improvement, Volume 2 Transgenic Approaches Editors Satbir Singh Gosal Shabir Hussain Wani School of Agricultural Biotechnology Mountain Research Centre Punjab Agricultural University for Field Crops, Khudwani Ludhiana, Punjab, India Sher-e-Kashmir University of Agricultural Sciences and Technology of Kashmir Srinagar, Jammu and Kashmir, India ISBN 978-3-319-90649-2 ISBN 978-3-319-90650-8 (eBook) https://doi.org/10.1007/978-3-319-90650-8 Library of Congress Control Number: 2018942727 © Springer International Publishing AG, part of Springer Nature 2018 This work is subject to copyright. All rights are reserved by the Publisher, whether the whole or part of the material is concerned, specifically the rights of translation, reprinting, reuse of illustrations, recitation, broadcasting, reproduction on microfilms or in any other physical way, and transmission or information storage and retrieval, electronic adaptation, computer software, or by similar or dissimilar methodology now known or hereafter developed. The use of general descriptive names, registered names, trademarks, service marks, etc. in this publication does not imply, even in the absence of a specific statement, that such names are exempt from the relevant protective laws and regulations and therefore free for general use. The publisher, the authors and the editors are safe to assume that the advice and information in this book are believed to be true and accurate at the date of publication. Neither the publisher nor the authors or the editors give a warranty, express or implied, with respect to the material contained herein or for any errors or omissions that may have been made. The publisher remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. Printed on acid-free paper This Springer imprint is published by the registered company Springer International Publishing AG part of Springer Nature. The registered company address is: Gewerbestrasse 11, 6330 Cham, Switzerland Professor Paul Christou Paul Christou obtained his Ph.D. in Organic Chemistry in 1980 at the University of London. He subsequently undertook postdoctoral research (1980–1982) at University College London in plant biochemistry with emphasis on the elucidation of the biosynthetic pathway of a number of irregular monoterpenoids useful in the perfumery and flavoring industries. In 1982, he was recruited by one of the first plant biotechnology companies in the USA, Agracetus Inc. (Madison, Wisconsin), as senior scientist with responsibilities for molecular and cellular plant biology. vi At Agracetus, he led a team that developed the first transgenic staple crop, soybean, currently being sold by Monsanto. His team also developed a facile genotype- independent method for rice transformation. In 1994, he moved to the John Innes Centre (JIC), Norwich, UK, as Head of Molecular Biotechnology Unit and Director of Tropical Maize and Rice Biotechnology Training Laboratory funded by the Rockefeller Foundation, USA. During his tenure at the JIC, he trained many Ph.D. students and hosted a number of postdoctoral fellows from 27 different countries in Asia, Africa, and Central and South America, funded by the Rockefeller Foundation, to pursue research in diverse aspects of rice and maize biotechnology. While at the JIC his team unraveled underpinning mechanisms controlling transgenic locus organization in cereal crops and pioneered a multigene transformation system useful in plant metabolic engineering. In 2001, he joined the Fraunhofer Institute of Molecular Biotechnology and Applied Ecology Schmallenberg-Aachen, Germany, as full professor. While at Fraunhofer he expanded his research to molecular pharming with emphasis on the production of recombinant proteins active against infectious diseases such as HIV. In 2004, he joined the University of Lleida, Spain, as an ICREA professor. He was the founding director of Agrotecnio Center, Lleida, Spain (2012), a position he held till 2015. He is currently the Editor in Chief of Molecular Breeding and Transgenic Research. He has mentored over 130 graduate students, and he is the senior vii author of over 250 scientific publications in the general area of plant biotechnology. He was awarded a European Research Council Advanced Grant (2009–2014) and a subsequent Proof of Concept Grant, also by the ERC. He has been a PI in 12 different EU-funded projects and has participated in 2 projects funded by the Bill & Melinda Gates Foundation, 1 currently in progress. He has given over 300 invited, plenary, or keynote lectures at international meetings. His current research focuses on metabolic engineering, genome editing and synthetic biology in cereals (rice and maize), production of pharmaceutical macromolecules in plants, and engineering novel agronomic traits in crops. He is active in training and technology transfer for developing country biotechnology, intellectual property issues, and regulatory and biosafety issues of transgenic crops, focusing on developing countries. He is also interested in science policy issues and strategic planning covering the interphase between fundamental and applied research. This book is dedicated to Prof. Paul Christou – A pioneer in particle gun technology for plant transformation. Foreword Agricultural production continues to face many challenges. Major constraints imposed by biotic and abiotic stresses on crop quality, yield, and productivity in conjunction with the urgent need to provide healthier and more nutritious food and animal feed in a durable and sustainable manner constitute one of the grand global challenges of our times. The challenge is even more severe in regions of the world which are less able to cope with increasing populations, changes in environmental and climatic conditions, pollution, and political and social instability. Agriculture has always been driven by technology. Its efficiency reflects game-changing discov- eries in biology and new developments in chemistry and engineering. We seek to increase food, feed, and fiber production while minimizing agriculture’s environ- mental footprint, and also to develop new products that branch into the chemical, pharmaceutical, and other industries. A key driver in this transition will be our abil- ity to control plant metabolism, development, biochemistry, and physiology holisti- cally, with more precision and predictability. Recent genome editing technologies are already beginning to make an impact on agriculture and are transforming the way improved and more resilient crop varieties are, and will continue to be created in the near future. However, it is important to recognize that older, more mature technologies still have the potential to play an enormous role in addressing the chal- lenges agriculture faces. Biotechnologies of Crop Improvement  – Transgenic Approaches, edited by Drs. Satbir Singh Gosal and Shabir Hussain Wani, provides an invaluable resource especially for students and younger practitioners who may be new to the field. The book comprises 16 chapters that address a number of impor- tant technologies all relying on transgenesis. The introductory chapter is a general overview of plant transformation methods. The chapter also provides examples of target traits under development. The subsequent four chapters have the common theme of gene silencing as a means to improve crops in a number of different ways. RNA interference and virus-induced gene silencing are being widely used to create plants which are more tolerant to biotic and abiotic stresses. These technologies have also been utilized to improve nutritional quality and overall physiology, devel- opment, and metabolism at the whole plant level. The next two chapters discuss antifungal plant defensins, small molecules with critical roles in plant immunity and ix x Foreword recent examples of plants with improved tolerance to salinity and drought. Specific chapters are devoted to the engineering of disease resistance in rice, an overview of the state of the art of sugarcane transformation and individual chapters on crops important for small-scale subsistence farmers such as pulses and millets. Tomato biotechnology is discussed in a dedicated chapter with emphasis on tolerance to environmental stresses and improvement of fruit quality traits. Eucalyptus genetic transformation is discussed in a separate chapter and target traits such as productiv- ity and quality for fiber production are highlighted. The importance of the enzyme glutamine synthetase in a number of different aspects of crop improvement is then discussed. This chapter highlights the importance of this enzyme in overall nitrogen metabolism in plants and by extension its role in improving nitrogen use efficiency a paramount challenge in agriculture. Its usefulness in creating new modes of herbi- cide tolerance in crops is discussed. The authors conclude the chapter by discussing inconsistent results reported in the literature concerning expression of glutamine synthase encoding genes in different plants. The penultimate chapter discusses the role of phytohormones in developing crops tolerant to environmental stresses. It focuses on underpinning mechanisms responsible for the role of phytohormones in plant metabolism. The last chapter provides a general historical overview of the development of transgenic crops, their adoption, and remaining challenges that still prevent such crops to reach those who need them the most, the small-scale subsis- tence farmers in the developing world. The last chapter also discusses next- generation precision engineering technologies using site-specific nucleases. Paul Christou University of Lleida Lleida, Spain Preface The combined use of recombinant DNA technology, gene-transfer methods, and tissue-culture techniques has led to the efficient transformation and production of transgenic plants in a wide variety of crop plants. In fact, transgenesis has emerged as an additional tool to carry out single-gene breeding or transgenic breeding of crops. The transgenic approach provides access to a larger gene pool, as the gene(s) may come from viruses, bacteria, fungi, insects, animals, human beings, unrelated plants, and even from chemical synthesis in the laboratory. Unlike conventional breeding, only the cloned gene(s) of agronomic importance is/are being introduced without co-transfer of undesirable genes/alleles from the donor parent. The recipi- ent genotype is least disturbed, which eliminates the need for repeated backcrosses. Various gene-transfer methods such as Agrobacterium, physicochemical uptake of DNA, liposome encapsulation, electroporation of protoplast, microinjection, DNA injection into intact plants, incubation of seeds with DNA, pollen tube pathway, laser microbeam, electroporation into tissues/embryos, silicon carbide fiber/whis- kers method, particle bombardment, and “in planta” transformation have been developed. Using different gene-transfer methods and strategies, transgenics carry- ing useful agronomic traits have been developed and released for commercial culti- vation. Attempts are being made to develop transgenic crop varieties resistant to abiotic stresses, such as drought, low and high temperature, salts, and heavy metals, and also to develop transgenic varieties possessing better nutrient-use efficiency and better keeping, nutritional and processing qualities. Genetically modified foods, such as tomato containing high lycopene, tomato with high flavonols as antioxi- dants, edible vaccines, are leading examples of genetically engineered crops. Several genes of agronomic importance have been isolated from various organisms; cloned and suitable constructs have been developed for plant transformation. Agrobacterium and “particle gun” methods have been refined and now being used for genetic transformation of a wide variety of field, fruit, vegetable, forest crops, and ornamental plant species. Transgenic crops such as maize, cotton, soybean, potato, tomato, papaya, and rice carrying mainly insect resistance, herbicide resis- tance, or both are now being commercially grown in several countries. xi

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During the past 15 years, cellular and molecular approaches have emerged as valuable adjuncts to supplement and complement conventional breeding methods for a wide variety of crop plants. Biotechnology increasingly plays a role in the creation, conservation, characterization and utilization of genet
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