Rajeev K Varshney FRS

Groundnut, an allotetraploid legume crop, serves as an important source of food, feed, and confectioneries worldwide. Translational genomics research has accelerated precision breeding efforts in groundnut using marker-assisted and genomic selection approaches. To support these efforts, three cost-effective and high-throughput genotyping panels have been developed to facilitate genetic studies and breeding applications. We developed high density genotyping array with 58K SNP markers which has been extensively used by global peanut research community. The first mid-density genotyping assay, comprises 5081 SNP markers, including 5000 highly informative SNPs with high polymorphism information content (PIC) derived from previously developed high-density ‘Axiom_Arachis’ array of 58,233 SNPs. An additional 81 SNPs associated with resilience and quality traits were incorporated for marker-assisted selection. This panel demonstrated robust performance, with PIC values ranging from 0.34 to 0.37 and explained 82.08% of genetic variance across 2573 genotypes from diverse sets of breeding populations. This panel holds promise for possible deployment in the identification of varietal seed mixture, genetic purity within gene bank germplasms and seed systems, foreground and background selection in backcross breeding programs, genomic selection, and sparse trait mapping studies in groundnut. The second panel, a mid-density array of 2500 SNP markers with an average density of 1 SNP per Mbp, is powerful tool for global groundnut breeding programmes. These SNPs were identified through whole-genome resequencing of 263 cultivated groundnut accessions representing diverse agronomic types and geographical regions. The panel includes 20-quality control (QC) and 72 associated markers for 8 key traits, namely leaf rust resistance, late leaf spot resistance, net blotch resistance, high oleic acid, seed weight, shelling percentage, fresh seed dormancy and blanchability. Designed through rigorous filtering processes, this panel is particularly suited for marker-assisted backcrossing, diversity analyses, and to study genetic purity for efficient germplasm management. Together, these genotyping panels provide versatile tools for advancing groundnut breeding programs, enabling precise and efficient genetic interventions for sustainable crop improvement.

In recent years, a truly impressive number of advances in genetics and genomics have greatly enhanced our understanding of structural and functional aspects of plant genomes. The complete genome sequences have become available for many crop species now. Due to advances in next generation sequencing technologies, it is possible to re-sequence mapping populations as well as germplasm collections in large numbers. In addition to routinely used linkage mapping and marker-assisted selection (MAS) approaches, several novel genetic and genomics tools/approaches such as genome-wide association studies (GWAS), marker-assisted recurrent selection (MARS), genomic selection (GS), functional genomics, etc. offer the possibilities to examine and utilize the structural as well as functional genetic variation in crop breeding. Holistic approach of enhancing the prediction of the phenotype from a genotype using genomics tools/approaches has been termed as 'genomics assisted breeding' (GAB) (Trends Plant Sci 10: 621 630). In fact, genomics assisted breeding has already shown its potential for crop improvement in several cereal species (Trends Biotech 24:490-499) and also in few legume species (Plant Breeding Rev 33:257-304). A critical assessment of the status and availability of genomic resources and genomics research in crop plant species and devising the strategies and approaches for effectively exploiting genomics research for crop improvement is the need of the hour (Nature Biotechnology 30:1172–1176). Genomics specialists having extensive experience in applying genomics in breeding from both public and private sectors share their experience and propose novel ideas in this workshop to use GAB in an efficient manner.

Chickpea (Cicer arietinum L.) is an important pulse crop grown in more than 50 countries across the globe especially in South Asia and Sub-Saharan Africa. During the last decade, genomic revolution empowered the chickpea community with large scale genomic resources for understanding the genetics of the trait and trait improvement using modern breeding approaches. Deploying the available genomic resources, we dissected important abiotic and biotic stresses that hinder chickpea production. The “QTL-hotspot” on CaLG04 explaining more than 58% phenotypic variation was introgressed into different elite backgrounds in India and Africa. Two high yielding and drought tolerant varieties, Geletu (in the background of ICCV 10) and BGM 10216 (in the background of Pusa 362) were released for commercial cultivation in Ethiopia and India respectively. In addition, MABC-WR-SA-1 a high yielding and Fusarium wilt resistant variety developed using marker assisted backcrossing approach was also release for comer cultivation in India. Following deciphering of the draft genome sequence of CDC Frontier variety, we re-sequenced >3000 chickpea germplasm accessions (that include global composite collection and 195 wild species accessions from primary, secondary and tertiary gene pools) at an average ~12X coverage. Large scale resequencing provided greater insights to genome-wide variations, the haplotype diversity, mutation burden, deleterious alleles, bottlenecks and selections sweeps during domestication. Extensive multi-location phenotyping data and the genome wide SNPs enabled us to identify genome-wide associations for agronomically important and > 40 nutritional quality traits. Furthermore, we are using these datasets for genomic prediction for developing climate resilient chickpea varieties.

Significant advances in recent years have enhanced our understanding of structural and functional aspects of plant genomes. Complete genome sequence has become available for some plant species, while draft genome sequences are being assembled for others. Advances in next-generation sequencing and high-throughput genotyping technologies have made it possible to not only genotype at high-density, but to even re-sequence the mapping populations or germplasm collections. Routine linkage mapping approaches are being transformed to genome-wide association studies (GWAS) for the identification of important marker-trait associations. Marker-assisted selection (MAS) has become an integral part of crop breeding programmes in developed countries/private sector, and in developing countries the use of MAS in breeding programmes has been applied to develop superior varieties/hybrids with better agronomic performance and enhanced resistance/tolerance to biotic/abiotic stresses. New breeding approaches such as marker-assisted back crossing (MABC), marker-assisted recurrent selection (MARS) and genomic selection (GS) are also being used for improving complex traits such as drought tolerance. This workshop is planned to invite genomics specialists with extensive experience in genomics-assisted breeding from both public and private sectors to share their experience and propose novel ideas to translate existing genomics knowledge toward the development of new varieties/hybrids. This workshop is expected to catalyze public sector breeding programmes in Asia and Africa to undertake/accelerate translational genomics that will ensure food security in developing countries.

Ascochyta blight, caused by Ascochyta rabiei, is globally the most destructive disease of chickpea. Chickpeas also suffer from emerging diseases, including seed rot and damping-off caused by Pythium ultimum. The objectives of this study were to detect molecular markers associated with resistance in chickpea to these two diseases. Plant materials included 177 chickpea recombinant inbred lines derived from an interspecific cross [C. reticulatum (PI 599072) x C. arietinum (FLIP 84-92C)] and 209 chickpea accessions representing an ICRISAT ‘mini-core’ collection. Disease screenings were conducted in a growth chamber (Pythium) or greenhouse (Ascochyta) in repeated experiments. A linkage map was developed for the C. reticulatum x C. arietinum population that included 1032 single nucleotide polymorphisms (SNPs) across eight linkage groups covering 965cM. Recombinant inbred lines were evaluated for reaction to seed rot and pre-emergence damping-off caused by metalaxyl resistant P. ultimum. A single significant QTL was detected on LG4 by composite interval mapping that explained 41.8 % of total phenotypic variation for disease reaction. Conversely, a genome wide association study (GWAS) approach was used to detect markers associated with resistance in the ICRISAT mini-core collection to Pythium seed rot and Ascochyta blight. A total of 302,902 single nucleotide polymorphisms (SNPs) equally distributed across the chickpea genome were examined with ADMIXTURE to determine population structure. Genomic regions were evaluated separately to establish marker-trait associations by employing FarmCPU and multiple loci mixed linear models (MMLM). For Pythium disease resistance, a total of 15 significant SNPs were detected and three candidate genes identified. One candidate gene and a total of 10 and 18 significant SNPs were detected for resistance to A. rabiei pathotype 1 (AR19) and pathotype 2 (AR628), respectively. SNPs and candidate genes identified in this study can be used for marker-assisted selection to develop superior chickpea varieties with improved disease resistance.

In order to facilitate low-cost and high-throughput genotyping for CGIAR centers and Partner institutes including National Agricultural Research Systems (NARS), a research initiative was established as “High Throughput Genotyping Project (HTPG)” led by ICRISAT with support from IRRI and CIMMYT through Excellence in Breeding (EiB) Platform. This project is funded by Bill and Melinda Gates Foundation (BMGF) to offer low-cost access to world-class genotyping services for CGIAR institutes and NARS partners. Under the HTPG service agreement, more than 40 public institutions globally (16+ species) listed as key users for the SNPLine genotyping platform. Primary objective of the project is to broker access of the latest genotyping platform to a wide user group at a reduced cost ($1.50 to $2.00 per sample) via sample aggregation. HTPG platform offers SNPs genotyping services in rice, wheat, maize, several millets, legumes and other crops, which are readily deployable in the breeding programmes. It also offers value added services, i.e. capacity building in high throughput sampling, digitization, barcoding and data interpretation support at no cost to partners. It helps in generation of economical, faster and quality SNP data for all genomics-related activities. The initiative also ensures that the latest molecular advances across the community brought under a single shared platform to facilitate effective and high impact collaboration. Lowering the genotyping cost will enable CGIAR, NARS and other public sector breeders to utilize marker-based selection in forward breeding and routine QC application in breeding pipeline to expedite the delivery of improved breeding products to farmers globally.

Grain legumes are source of high-quality food and feed and their integration in the cropping systems provide multiple benefits for agriculture sustainability. They are being considered important to ensure food and nutritional security in the face of climate change. Diseases and insect pests are the main constraints in the quantity and quality of yield. This presentation focuses on chickpea (Cicer arietinum L.) and pigeonpea (Cajanus cajan L.), which are mandate crops of ICRISAT and globally grown on 21.6 million ha, largely in the developing countries of Asia and Africa. The production and productivity of chickpea is severely constrained by diseases such as Fusarium wilt (FW, Fusarium oxysporum f sp ciceris), dry root rot (DRR, Rhizoctonia bataticola), Ascochyta blight (AB, Ascochyta rabiei) and Botrytis gray mold (BGM, Botrytis cinerea). In pigeonpea, Fusarium wilt (FW, Fusarium udum) and sterility mosaic disease (SMD) caused by pigeonpea sterility mosaic virus (PPSMV) are the most important diseases, while Phytophthora blight (PB, Phytophthora cajani) is an emerging important disease. Pod borer [Helicoverpa armigera (Hubner)] is the most important insect-pest of both the legumes. In addition, spotted pod borer [(Maruca vitrata (Geyer)] is also important in pigeonpea. Several varieties with high resistance to FW and AB have been developed in chickpea and FW and SMD in pigeonpea. Only moderate level of resistance is available for resistance to the remaining diseases and pod borers in the germplasm of cultivated species. Comparatively, higher levels of resistance have been observed for some of these diseases and pod borers in wild species and are being exploited in breeding programs. Transgenic resistance using insecticidal genes has been developed to achieve high level of resistance to pod borer in both the legumes. Numerous genes/quantitative trait loci (QTL) conferring resistance to key diseases have been mapped and markers linked to some of these have been validated. Use of novel sources of resistance and novel breeding techniques (marker-assisted selection, speed breeding) are being used to accelerate development of improved varieties with enhanced resistance to diseases and insect pests. Availability of such varieties will improve yield stability and production of these grain legumes and contribute to food and nutritional security and sustainable food production.

Pearl millet (Pennisetum glaucum L.) is the sixth most important cereal crop globally, predominantly grown for food and forage in arid and semi-arid tropical regions. Under climate change, pearl millet will face more adverse climatic conditions, particularly due to drought and heat stress and newly emerging disease, blast. Crop wild relatives (CWR) are the reservoir of valuable genes for tolerance/resistance to various abiotic/biotic stresses. Primary genepool species Pennisetum glaucum subsp. violaceum evolved in hot and dry conditions of Sahel region in Africa, grows in even more arid regions than the cultigen, and thus possess higher levels of drought and heat tolerance. Four pre-breeding populations were developed using wild Pennisetum violaceum accessions and cultivated pearl millet genotypes following advanced backcross approach. These populations were evaluated for flowering-stage heat stress during 2018 summer season across three locations, Agra in Uttar Pradesh, and SK Nagar and Tharad in Gujarat in India. These populations were also evaluated for terminal drought during 2018 rainy season at two locations, Hisar and Bawal. Promising ILs having improved heat and drought tolerance were identified. These populations were also evaluated under LeasyScan for the canopy-related parameters, as well as at LysiField facility for the traits related to water-use and water- use efficiency under well-watered and water-stress conditions. Further, screening of promising drought and/or heat tolerant ILs for five diverse pathotypes, Pg 45, Pg 138, Pg 186, Pg 204 and Pg 232 of blast resulted in the identification of resistant ILs. Preliminary screening of three populations for Striga hermonthica in Niger resulted in the identification of several ILs having improved Striga resistance. Re-screening of the material will be done during 2019 rainy season to confirm the results. Utilization of these promising ILs in breeding programs will assist in developing new varieties/hybrids with improved tolerance to important biotic/abiotic stresses.

Although crop improvement programs have made excellent progress in enhancing crop productivity and production, there is still a huge scope to fill the yield gap for majority of crops in dryland areas. Genomics-assisted breeding can help enhancing crop productivity as well as nutrition in these crops. However, until recently, majority of the dryland crops have remained untouched with genomics revolution. Two key reasons for this situation include engagement of only few institutes and availability of limited resources at international level for research and development in these crops. With an objective to address these issues, the Center of Excellence in Genomics and Systems Biology (CEGSB) at International Crops Research Institute for the Semi-Arid Tropics (ICRISAT) floated several multi-institutional consortia. As a result of collaborative efforts from such strong partnership, a large number of genomic resources including genome assemblies for 9 crops have been developed and several improved lines have been developed through molecular breeding. In summary, translational genomics approach has transformed the so-called ‘orphan crops’ to ‘genomic resources-rich crops’ and contributed to develop several improved lines in some dryland crops.

Legumes are second to cereals in agricultural production accounting for approximately 27% of the world’s primary production. Grain legumes alone contribute to about one-third of the human dietary protein. Chickpea is one of the world’s largest cultivated food legume and is an excellent source of high-quality protein to the human diet. Plant growth and development are controlled by programmed expression of a suite of genes at the given time, stage and tissue. Understanding how the underlying genome sequence translates into specific plant phenotypes at key developmental stages, information on gene expression patterns is crucial. Here we present a comprehensive Cicer arietinum Gene Expression Atlas (CaGEA) across the plant developmental stages and organs covering the entire life cycle of chickpea. One of the widely used drought tolerant cultivar, ICC 4958 has been used to generate RNA-Seq data from 27 samples at five major developmental stages of the plant. A total of 816 million raw reads were generated and of these, 794 million filtered reads after QC were subjected to downstream analysis. A total of 15,947 unique number of differentially expressed genes across different pairwise tissue combinations were identified. Significant differences in gene expression patterns contributing in the process of flowering, nodulation, seed and root development were inferred in this study. Furthermore, four functionally relevant candidate genes on the effect of “QTL-hotspot” for drought tolerance in chickpea were identified. The gene expression atlas of chickpea would not only provide a global view of gene expression patterns in all major tissues and organ systems but also will serve as valuable resource for functional genomics and accelerate gene discovery in legumes.