Genotyping by Sequencing for Crop Improvement. Группа авторов

Vijayan, ICAR – National Institute for Biotechnology, New Delhi, India

      Dhiraj Lalji Wasule, Vasantrao Naik College of Agricultural Biotechnology, Dr. Panjabrao Deshmukh Krishi Vidyapeeth, Akola, Maharashtra, India

      Himanshu Yadav, Department of Agriculture Biotechnology, National Agri‐Food Biotechnology Institute (NABI), Mohali, Punjab, India

Preface

      Recent advances in sequencing technology and computational resources have accelerated genomics and translational research in crop science. The technological advances have provided many opportunities in genomics‐assisted plant breeding to address issues related to food security. Among the several applications, genotyping‐by‐sequencing (GBS) technology has evolved as one of the frontier areas facilitating high‐throughput plant genotyping. The GBS approaches have proved effective for the utilization in genotyping‐based applications like quantitative trait loci (QTL) mapping, genome‐wide association study (GWAS), genomic selection (GS), and marker‐assisted breeding (MAB). Considering the current affairs in plant breeding, we decided to compile the advances in GBS methods, statistical approaches to analyze the GBS data, and its applications including QTL mapping, GWAS, and GS in crop improvement.

      Presently, the food produced around the world is adequate for the existing population. However, the constantly increasing population mounting pressure on a food production system. Hence efficient utilization of technological advances and existing knowledge is essential to enhance food production to match the growing food demand. In this direction, most of the countries around the globe have adopted advanced genomic methodologies to breed superior plant genotypes. Among such technological advances, the high‐throughput genotyping using GBS has shown promising results in different crop plants. The GBS has predominantly been used for germplasm evaluation, evolutionary studies, development of dense linkage map, QTL mapping, GWAS, GS, and MAB. The cost‐effectiveness and whole‐genome coverage make GBS more reliable than other next‐generation sequencing (NGS) techniques.

This book describes advanced molecular markers, high‐throughput genotyping platforms, whole‐genome resequencing (WGR), QTL mapping using advanced mapping populations, analytical pipelines for the GBS analysis, advances in GWAS, advances in GS, application of GBS, GWAS, and GS in different crop plants. The different marker types including traditional and advanced markers used in plant genotyping have been presented in great detail. DNA extraction directly from seeds without germination can save time and effort. Several modified and crop‐specific nondestructive seed DNA extraction protocols have been compiled and presented. Many advanced genotyping platforms are now available which cater to specific research purposes because of the differences in terms of reaction chemistry involved, cost, method of signal detection, and flexibility in the protocols. Such advanced platforms along with their principles have been discussed. The WGR methodology and available resources have been covered in detail. The WGR has emerged as a powerful method to identify genetic variation among individuals. The recent advancement in WGR includes pool‐Seq which provides an alternative to individual sequencing and a cost‐effective method for GWAS. Compared to biparental populations the multi‐parental population provides an opportunity to interrogate multiple alleles and to provide an increased level of recombination and mapping resolution of QTLs. The use of such improved populations in the era of high‐throughput genotyping has been presented in one of the chapters. The dedicated section focused on the basic principle of GWAS, the efficiency of different markers, candidate gene identification, meta‐GWAS, and statistical methods involved in GWAS analysis has been included. For genetic mapping, and marker‐assisted selection, rapid and quality DNA isolation is mandatory to accelerate the whole process. A focused section about GS has been included which gives an account of the basic concept, advances, applicability, and challenges of GS. Similarly, a separate chapter is included which discusses the analytical pipelines used for GBS data. Application of technologies such as GBS, GWAS, and GS in different crop categories like cereals, pulses, oilseeds, and commercial crops has been discussed in different chapters.

      Here, we have tried to compile basic aspects and recent advances in GBS, GWAS, and GS in plant breeding. We believe that the book will be helpful to researchers and scientists to understand and plan future experiments. This book will enable plant scientists to explore GBS application more efficiently for basic research as well as applied aspects in various crops improvement projects.

      Editors

      Dr. Humira Sonah

      Dr. Vinod Goyal

      Dr. S. M. ShivarajDr. R. K. Deshmukh

1 Molecular Marker Techniques and Recent Advancements

       Dharminder Bhatia and Gagandeep Singh Bajwa

       Department of Plant Breeding and Genetics, Punjab Agricultural University, Ludhiana, Punjab, India

1.1 Introduction

      Plant selection and systematic breeding efforts led to the development of present‐day improved cultivars of crop plants. From a historical perspective, increased crop yield is the result of genetic improvement (Fehr 1984). Markers play an important role in the selection of traits of interest. Markers can be morphological, biochemical, or molecular in nature. Morphological markers are visual phenotypic characters such as growth habit of the plant, seed shape, seed color, flower color etc. Biochemical markers are the isozyme‐based markers characterized by variation in molecular form of enzyme showing a difference in mobility on an electrophoresis gel. Very few morphological and biochemical markers are available in plants, and they are influenced by developmental stage and environmental factors. Since a large number of economically important traits are quantitative in nature, which are affected by both genetic and environmental factors, the morphological and biochemical markers‐based selection of traits may not be much reliable. The subsequent discovery of abundantly available DNA‐based markers made possible the selection of almost any trait of interest. DNA‐based markers are not affected by the environment. Besides, these markers are highly reproducible across labs and show high polymorphism to distinguish between two genetically different individuals or species.

In the last four decades, DNA‐based molecular marker technology has witnessed several advances from low throughput hybridization‐based markers to high‐throughput sequencing‐based markers. These advances have been possible due to critical discoveries such as polymerase chain reaction (PCR) (Mullis et al. 1986), Sanger sequencing method (Sanger et al. 1977), automation of Sanger sequencing (Shendure et al. 2011), next‐generation sequencing (NGS) technologies (Mardis 2008), and development of bioinformatics tools. This chapter will briefly discuss different types of molecular markers while particularly focusing on recent developments in molecular marker technologies. These developments have expedited the mapping and cloning of several loci governing important traits, precise trait selection, and transfer into elite germplasm.

1.2 What is a Molecular Marker?

      DNA or molecular marker is a fragment of the DNA that is associated with a particular trait in an individual. These molecular markers aid in determining the location of genes that control key traits.

      Generally, molecular markers do not represent the gene of interest but act as “flags” or “signs.” Similar to genes, all the molecular markers occupy a specific position within the chromosomes. Molecular markers located close to genes (i.e. tightly linked) are referred to as “gene tags.”

      DNA‐based molecular markers are the most widely used markers predominantly due to their abundance. They arise from different classes of DNA mutations such as substitution mutations (point mutations), rearrangements (insertions or deletions), or errors in replication of tandemly repeated DNA. These markers are selectively neutral because they are usually located in noncoding regions of DNA. Unlike morphological and biochemical markers, DNA markers are practically unlimited