Kalyani Srinivasan

Comparison of SSR and SNP markers in estimation of genetic diversity and population structure of Indian rice varieties.

Simple sequence repeat (SSR) and Single Nucleotide Polymorphic (SNP), the two most robust markers for identifying rice varieties were compared for assessment of genetic diversity and population structure. Total 375 varieties of rice from various regions of India archived at the Indian National GeneBank, NBPGR, New Delhi, were analyzed using thirty six genetic markers, each of hypervariable SSR (HvSSR) and SNP which were distributed across 12 rice chromosomes. A total of 80 alleles were amplified with the SSR markers with an average of 2.22 alleles per locus whereas, 72 alleles were amplified with SNP markers. Polymorphic information content (PIC) values for HvSSR ranged from 0.04 to 0.5 with an average of 0.25. In the case of SNP markers, PIC values ranged from 0.03 to 0.37 with an average of 0.23. Genetic relatedness among the varieties was studied; utilizing an unrooted tree all the genotypes were grouped into three major clusters with both SSR and SNP markers. Analysis of molecular variance (AMOVA) indicated that maximum diversity was partitioned between and within individual level but not between populations. Principal coordinate analysis (PCoA) with SSR markers showed that genotypes were uniformly distributed across the two axes with 13.33% of cumulative variation whereas, in case of SNP markers varieties were grouped into three broad groups across two axes with 45.20% of cumulative variation. Population structure were tested using K values from 1 to 20, but there was no clear population structure, therefore Ln(PD) derived Δk was plotted against the K to determine the number of populations. In case of SSR maximum Δk was at K=5 whereas, in case of SNP maximum Δk was found at K=15, suggesting that resolution of population was higher with SNP markers, but SSR were more efficient for diversity analysis.

Analysis of Genetic Diversity and Population Structure of Rice Germplasm from North-Eastern Region of India and Development of a Core Germplasm Set

The North-Eastern region (NER) of India, comprising of Arunachal Pradesh, Assam, Manipur, Meghalaya, Mizoram, Nagaland and Tripura, is a hot spot for genetic diversity and the most probable origin of rice. North-east rice collections are known to possess various agronomically important traits like biotic and abiotic stress tolerance, unique grain and cooking quality. The genetic diversity and associated population structure of 6,984 rice accessions, originating from NER, were assessed using 36 genome wide unlinked single nucleotide polymorphism (SNP) markers distributed across the 12 rice chromosomes. All of the 36 SNP loci were polymorphic and bi-allelic, contained five types of base substitutions and together produced nine types of alleles. The polymorphic information content (PIC) ranged from 0.004 for Tripura to 0.375 for Manipur and major allele frequency ranged from 0.50 for Assam to 0.99 for Tripura. Heterozygosity ranged from 0.002 in Nagaland to 0.42 in Mizoram and gene diversity ranged from 0.006 in Arunachal Pradesh to 0.50 in Manipur. The genetic relatedness among the rice accessions was evaluated using an unrooted phylogenetic tree analysis, which grouped all accessions into three major clusters. For determining population structure, populations K = 1 to K = 20 were tested and population K = 3 was present in all the states, with the exception of Meghalaya and Manipur where, K = 5 and K = 4 populations were present, respectively. Principal Coordinate Analysis (PCoA) showed that accessions were distributed according to their population structure. AMOVA analysis showed that, maximum diversity was partitioned at the individual accession level (73% for Nagaland, 58% for Arunachal Pradesh and 57% for Tripura). Using POWERCORE software, a core set of 701 accessions was obtained, which accounted for approximately 10% of the total NE India collections, representing 99.9% of the allelic diversity. The rice core set developed will be a valuable resource for future genomic studies and crop improvement strategies.

Evaluation of 19,460 Wheat Accessions Conserved in the Indian National Genebank to Identify New Sources of Resistance to Rust and Spot Blotch Diseases

A comprehensive germplasm evaluation study of wheat accessions conserved in the Indian National Genebank was conducted to identify sources of rust and spot blotch resistance. Genebank accessions comprising three species of wheat–Triticum aestivum, T. durum and T. dicoccum were screened sequentially at multiple disease hotspots, during the 2011–14 crop seasons, carrying only resistant accessions to the next step of evaluation. Wheat accessions which were found to be resistant in the field were then assayed for seedling resistance and profiled using molecular markers. In the primary evaluation, 19,460 accessions were screened at Wellington (Tamil Nadu), a hotspot for wheat rusts. We identified 4925 accessions to be resistant and these were further evaluated at Gurdaspur (Punjab), a hotspot for stripe rust and at Cooch Behar (West Bengal), a hotspot for spot blotch. The second round evaluation identified 498 accessions potentially resistant to multiple rusts and 868 accessions potentially resistant to spot blotch. Evaluation of rust resistant accessions for seedling resistance against seven virulent pathotypes of three rusts under artificial epiphytotic conditions identified 137 accessions potentially resistant to multiple rusts. Molecular analysis to identify different combinations of genetic loci imparting resistance to leaf rust, stem rust, stripe rust and spot blotch using linked molecular markers, identified 45 wheat accessions containing known resistance genes against all three rusts as well as a QTL for spot blotch resistance. The resistant germplasm accessions, particularly against stripe rust, identified in this study can be excellent potential candidates to be employed for breeding resistance into the background of high yielding wheat cultivars through conventional or molecular breeding approaches, and are expected to contribute toward food security at national and global levels.

Studies on the seasonal abundance and control of okra stemfly, Melanagromyza hibisci Spencer (Diptera: Agromyzidae) in India

Incidence of okra stemfly, Melanagromyza hibisci Spencer in India was high (40–60%) in winter months, i.e. during October, November, December and January sowings. Mean maximum and minimum temperature that prevailed 30 days after sowing had significant negative correlation with percentage main stem damage. Carbofuran, phorate, quinalphos and aldicarb, applied at 1 kg per ha reduced damage to main stem at the seedling stage but could not control subsequent damage to petioles. Eurytoma sp., a parasitoid on M. hibisci, had a high density-dependent relationship with its host, and played a significant role in keeping it under control.RésuméL’influence de la mouche okra [Melanagromyza hibisci Spencer] sur les tiges des semailles pendant les mois d’hiver c’est-à-dire en octobre, novembre, décembre et janvier est ‘élevé’ de 40–60% de plantes parasitées. La température moyenne maximale et minimale qui prevant 30 jours après les semilles a montre une corrélation négative significative par rapport au pourcentage des dommages sur la tige centrale carbofuran, quinalphos, phorate et aldicarb répandus à raision d’un kilo/hectare, a réduit les dommages sur la tige centrale de la jeune poussé mais n’a en aucune influence sur ceux des pétioles. Eurytoma sp. un parasite de la mouche M. hibisci, très dépendante de son hôte, à joue un rôle significatif pour le contrôler.

Non-destructive Method of Seed Vigour Testing of Traditional Rice (Oryza sativa L.) Varieties Conserved in Genebank under Medium-term Storage Conditions

Plant genetic resources are commonly conserved in the form of seeds in genebanks for their present and future use in crop improvement programmes. It is commonly understood that the loss in vigour, precedes loss in viability during storage. However, vigour tests cannot be conducted routinely for a large number of varieties processed for conservation and monitoring seed quality in genebanks. Hence, rapid and non-destructive tests will be immensely useful to provide additional information during monitoring of seed samples. The seeds of 31 traditional varieties of rice, conserved in medium-term storage (MTS) module (4°C and 35% relative humidity) at ICAR-National Bureau of Plant Genetic Resources, New Delhi, for past 10 years, were used to assess the seed quality by recording the germination percentage, mean germination time, seedling length and vigour index I, electrical conductivity, potassium (K+) and sodium (Na+) ion concentration in leachates. Results of the study clearly confirmed the fact that K+, Na+ ion concentrations in leachate and K+/Na+ ratio can be used as reliable and non-destructive method of vigour tests for assessing the quality of rice seeds. Several varieties with similar high percentage of initial germination deteriorated drastically leading to highly variable germination percentages after 10 years in MTS, indicating genotype-specificity.

Development of Core Set of Wheat ( Triticum spp. ) Germplasm Conserved in the National Genebank in India

Plant genetic resources, the source of genetic diversity provides a broad genetic foundation for plant breeding and genetic research, however, large germplasm resources are difficult to preserve, evaluate and use. Construction of core and mini core collections is an efficient method for managing genetic resources and undertaking intensive surveys of natural variation, including the phenotyping of complex traits and genotyping of DNA polymorphisms allowing more efficient utilization of genetic resources. A mega characterization and evaluation programme of the entire cultivated gene pool of wheat conserved in the National Genebank, India was undertaken. Wheat accessions with limited seed quantity, were multiplied in the off-season nursery at IARI Regional Station, Wellington during rainy season 2011 and the entire set of 22,469 wheat accessions were characterized and evaluated at CCS HAU, Hisar, Haryana during winter season 2011–12 for 34 characters including 22 highly heritable qualitative, and 12 quantitative parameters. The core sets were developed using PowerCore Software with stepwise approach and grouping method and validated using Shannon-Diversity Index and summary statistics. Based on Shannon-Diversity index, PowerCore with stepwise approach was found better than PowerCore with grouping. The core set included 2,208 accessions comprising 1,770 T. aestivum, 386 T. durum, and 52 T. dicoccum accessions as a representative of the total diversity recorded in the wheat germplasm. The core set developed will be further validated at different agro-climatic conditions and will be utilized for development of mini core set to enhance the utilization by wheat researchers and development of climate resilient improved varieties.