K. K. Vinod

Molecular evaluation of genetic diversity and association studies in rice (Oryza sativa L.).

In the present study, we tested rice genotypes that included un(der)exploited landraces of Tamil Nadu along with indica and japonica test cultivars to ascertain their genetic diversity structure. Highly polymorphic microsatellite markers were used for generating marker segregation data. A novel measure, allele discrimination index, was used to determine subpopulation differentiation power of each marker. Phenotypic data were collected for yield and component traits. Pattern of molecular differentiation separated indica and japonica genotypes; indica genotypes had two subpopulations within. Landraces were found to have indica genome, but formed a separate subgroup with low linkage disequilibrium. The landraces further separated into distinct group in both hierarchical clustering analysis using neighbour-joining method as well as in the model based population structure analysis. Japonica and the remaining indica cultivars formed two other distinct groups. Linkage disequilibrium observed in the whole population was considerably reduced in subpopulations. Low linkage disequilibrium of landforms suggests their narrow adaptation in local geographical niche. Many population specific alleles could be identified particularly for japonica cultivars and landraces. Association analysis revealed nine marker–trait associations with three agronomic traits, of which 67% were previously reported. Although the testing landraces together with known cultivars had permitted genome-wide association mapping, the experiment offers scope to study more landraces collected from the entire geographical region for drawing more reliable information.

Molecular breeding for the development of multiple disease resistance in Basmati rice

Marker assisted backcross breeding for combining three resistance genes (xa13 and Xa21 for Bacterial Blight, Pi54 for blast) and a major QTL (qSBR11-1 for resistance to Sheath blight) in Basmati rice.

QTL and QTL × Environment Effects on Agronomic and Nitrogen Acquisition Traits in Rice

Agricultural environments deteriorate due to excess nitrogen application. Breeding for low nitrogen responsive genotypes can reduce soil nitrogen input. Rice genotypes respond variably to soil available nitrogen. The present study attempted quantification of genotype x nitrogen level interaction and mapping of quantitative trait loci (QTLs) associated with nitrogen use efficiency (NUE) and other associated agronomic traits. Twelve parameters were observed across a set of 82 double haploid (DH) lines derived from IR64/Azucena. Three nitrogen regimes namely, native (0 kg/ha; no nitrogen applied), optimum (100 kg/ha) and high (200 kg/ha) replicated thrice were the environments. The parents and DH lines were significantly varying for all traits under different nitrogen regimes. All traits except plant height recorded significant genotype x environment interaction. Individual plant yield was positively correlated with nitrogen use efficiency and nitrogen uptake. Sixteen QTLs were detected by composite interval mapping. Eleven QTLs showed significant QTL x environment interactions. On chromosome 3, seven QTLs were detected associated with nitrogen use, plant yield and associated traits. A QTL region between markers RZ678, RZ574 and RZ284 was associated with nitrogen use and yield. This chromosomal region was enriched with expressed gene sequences of known key nitrogen assimilation genes.

Trait identification and QTL validation for reproductive stage drought resistance in rice using selective genotyping of near flowering RILs

Drought resistance is becoming an indispensable character for rice improvement due to the dwindling global water resources. Genetic improvement for drought resistance is achieved through physiological dissection and genetic analysis of independent component traits associated with crop productivity under stress. A subset mapping population of 93 near flowering recombinant inbred lines with uniform phenology was constituted for genetic analysis of reproductive stage drought resistance. The population was phenotyped for 22 physio-morphological traits under two contrasting water regimes imposed at reproductive stage. Broad sense heritabilities of morphological traits were lower under stress than irrigated. Predominant association of plant height, panicle exsertion and harvest index with grain yield were observed under stress. The sustenance of panicle exsertion through maintaining growth during moisture stress was found as a significant trait associated with the grain yield through minimizing spikelet sterility. Selective genotyping was carried out with 23 polymorphic microsatellite markers of the established target genomic regions for drought resistance. The study validated the association of a QTL region on the long arm of chromosome 1 with plant height, panicle length, panicle exsertion, biological yield and stomatal conductance under stress. This region, flanked by markers RM246 and RM315, was known to possess the semi-dwarf gene, sd-1. Role of another major interval lying between RM256 and RM149 on chromosome 8 in defining the drought resistance could be established through identification of QTLs associated with leaf rolling, panicle exsertion, plant height, panicle length, senescence and biological yield under moisture stress condition. Few other QTLs were also identified.

Improvement of Basmati rice varieties for resistance to blast and bacterial blight diseases using marker assisted backcross breeding.

Marker assisted backcross breeding was employed to incorporate the blast resistance genes, Pi2 and Pi54 and bacterial blight (BB) resistance genes xa13 and Xa21 into the genetic background of Pusa Basmati 1121 (PB1121) and Pusa Basmati 6. Foreground selection for target gene(s) was followed by arduous phenotypic and background selection which fast-tracked the recovery of recurrent parent genome (RPG) to an extent of 95.8% in one of the near-isogenic lines (NILs) namely, Pusa 1728-23-33-31-56, which also showed high degree of resemblance to recurrent parent, PB6 in phenotype. The phenotypic selection prior to background selection provided an additional opportunity for identifying the novel recombinants viz., Pusa 1884-9-12-14 and Pusa 1884-3-9-175, superior to parental lines in terms of early maturity, higher yield and improved quality parameters. There was no significant difference between the RPG recovery estimated based on SSR or SNP markers, however, the panel of SNPs markers was considered as the better choice for background selection as it provided better genome coverage and included SNPs in the genic regions. Multi-location evaluation of NILs depicted their stable and high mean performance in comparison to the respective recurrent parents. The Pi2+Pi54 carrying NILs were effective in combating a pan-India panel of Magnaporthe oryzae isolates with high level of field resistance in northern, eastern and southern parts of India. Alongside, the PB1121-NILs and PB6-NILs carrying BB resistance genes xa13+Xa21 were resistant against Xanthomonas oryzae pv. oryzae races of north-western, southern and eastern parts of the country. Three of NILs developed in this study, have been promoted to final stage of testing during the ​Kharif 2015 in the Indian National Basmati Trial.

Approaches towards nitrogen- and phosphorus-efficient rice

Crop tolerance to lowered availability of nutrients is a major breeding objective in rice. Current understanding of complex genetic control of N and P utilisation is being converged towards precision breeding such as marker assisted breeding for nutrient efficient varieties.

Assessment of genetic diversity in Indian rice germplasm (Oryza sativa L.): use of random versus trait-linked microsatellite markers

Assessment of genetic diversity in a crop germplasm is a vital part of plant breeding. DNA markers such as microsatellite or simple sequence repeat markers have been widely used to estimate the genetic diversity in rice. The present study was carried out to decipher the pattern of genetic diversity in terms of both phenotypic and genotypic variability, and to assess the efficiency of random vis-à-vis QTL linked/gene based simple sequence repeat markers in diversity estimation. A set of 88 rice accessions that included landraces, farmer’s varieties and popular Basmati lines were evaluated for agronomic traits and molecular diversity. The random set of SSR markers included 50 diversity panel markers developed under IRRI’s Generation Challenge Programme (GCP) and the trait-linked/gene based markers comprised of 50 SSR markers reportedly linked to yield and related components. For agronomic traits, significant variability was observed, ranging between the maximum for grains/panicle and the minimum for panicle length. The molecular diversity based grouping indicated that varieties from a common centre were genetically similar, with few exceptions. The trait-linked markers gave an average genetic dissimilarity of 0.45 as against that of 0.37 by random markers, along with an average polymorphic information constant value of 0.48 and 0.41 respectively. The correlation between the kinship matrix generated by trait-linked markers and the phenotype based distance matrix (0.29) was higher than that of random markers (0.19). This establishes the robustness of trait-linked markers over random markers in estimating genetic diversity of rice germplasm.

Marker-aided Incorporation of Xa38 , a Novel Bacterial Blight Resistance Gene, in PB1121 and Comparison of its Resistance Spectrum with xa13 + Xa21

Basmati rice is preferred internationally because of its appealing taste, mouth feel and aroma. Pusa Basmati 1121 (PB1121) is a widely grown variety known for its excellent grain and cooking quality in the international and domestic market. It contributes approximately USD 3 billion to India’s forex earning annually by being the most traded variety. However, PB1121 is highly susceptible to bacterial blight (BB) disease. A novel BB resistance gene Xa38 was incorporated in PB1121 from donor parent PR114-Xa38 using a modified marker-assisted backcross breeding (MABB) scheme. Phenotypic selection prior to background selection was instrumental in identifying the novel recombinants with maximum recovery of recurrent parent phenome. The strategy was effective in delimiting the linkage drag to <0.5 mb upstream and <1.9 mb downstream of Xa38 with recurrent parent genome recovery upto 96.9% in the developed NILs. The NILs of PB1121 carrying Xa38 were compared with PB1121 NILs carrying xa13 + Xa21 (developed earlier in our lab) for their resistance to BB. Both NILs showed resistance against the Xoo races 1, 2, 3 and 6. Additionally, Xa38 also resisted Xoo race 5 to which xa13 + Xa21 was susceptible. The PB1121 NILs carrying Xa38 gene will provide effective control of BB in the Basmati growing region.

Improving Salt Tolerance in Rice: Looking Beyond the Conventional

Several factors in the intensive cropping system have played significant role in deteriorating soil health in general. soil salinization is one of the major issues threatening crop productivity in major irrigated rice growing areas of the world. Salinity is a serious issue in rice, the crop that feeds half the world, since it is sensitive to salt accumulation. With the world population growing incessantly, there is an urgent need to increase rice productivity especially in salinized lands as well as to reutilize lands that are rendered unproductive due to salt accumulation. It is therefore essential to develop varieties that are phenologically capable of sustaining excess salt throughout its life span and produce higher yield. Although there is sufficient variability in rice germplasm for salt tolerance, conventional breeding has been far less fruitful in addressing this complex problem. With the deeper understanding of the intricate mechanisms of salt tolerance and the array of genes and useable quantitative trait loci that are being discovered, the breeding scenario towards salt tolerant rice is poised to take a more productive turn in near future. This chapter outlines the latest developments in rice breeding towards salt tolerance through employment of modern molecular techniques in conjunction with the conventional breeding approaches.

Tracing QTLs for Leaf Blast Resistance and Agronomic Performance of Finger Millet (Eleusine coracana (L.) Gaertn.) Genotypes through Association Mapping and in silico Comparative Genomics Analyses

Finger millet is one of the small millets with high nutritive value. This crop is vulnerable to blast disease caused by Pyricularia grisea, which occurs annually during rainy and winter seasons. Leaf blast occurs at early crop stage and is highly damaging. Mapping of resistance genes and other quantitative trait loci (QTLs) for agronomic performance can be of great use for improving finger millet genotypes. Evaluation of one hundred and twenty-eight finger millet genotypes in natural field conditions revealed that leaf blast caused severe setback on agronomic performance for susceptible genotypes, most significant traits being plant height and root length. Plant height was reduced under disease severity while root length was increased. Among the genotypes, IE4795 showed superior response in terms of both disease resistance and better agronomic performance. A total of seven unambiguous QTLs were found to be associated with various agronomic traits including leaf blast resistance by association mapping analysis. The markers, UGEP101 and UGEP95, were strongly associated with blast resistance. UGEP98 was associated with tiller number and UGEP9 was associated with root length and seed yield. Cross species validation of markers revealed that 12 candidate genes were associated with 8 QTLs in the genomes of grass species such as rice, foxtail millet, maize, Brachypodium stacei, B. distachyon, Panicum hallii and switchgrass. Several candidate genes were found proximal to orthologous sequences of the identified QTLs such as 1,4-β-glucanase for leaf blast resistance, cytokinin dehydrogenase (CKX) for tiller production, calmodulin (CaM) binding protein for seed yield and pectin methylesterase inhibitor (PMEI) for root growth and development. Most of these QTLs and their putatively associated candidate genes are reported for first time in finger millet. On validation, these novel QTLs may be utilized in future for marker assisted breeding for the development of fungal resistant and high yielding varieties of finger millet.