Integrated Physiological, Biochemical, and Molecular Analysis Identifies Important Traits and Mechanisms Associated with Differential Response of Rice Genotypes to Elevated Temperature

In changing climate, heat stress caused by high temperature poses a serious threat to rice cultivation. A multiple organizational analysis at physiological, biochemical and molecular level is required to fully understand the impact of elevated temperature in rice. This study was aimed at deciphering the elevated temperature response in eleven popular and mega rice cultivars widely grown in India. Physiological and biochemical traits specifically membrane thermostability (MTS), antioxidants, and photosynthesis were studied at vegetative and reproductive phases which were used to establish a correlation with grain yield under stress. Several useful traits in different genotypes were identified which will be important resource to develop high temperature tolerant rice cultivars. Interestingly, Nagina22 emerged as best performer in terms of yield as well as expression of physiological and biochemical traits at elevated temperature. It showed lesser relative injury, lesser reduction in chlorophyll content, increased super oxide dismutase, catalase and peroxidase activity, lesser reduction in net photosynthetic rate (PN), high transpiration rate (E) and other photosynthetic/ fluorescence parameters contributing to least reduction in spikelet fertility and grain yield at elevated temperature. Further, expression of 14 genes including heat shock transcription factors and heat shock proteins was analyzed in Nagina22 (tolerant) and Vandana (susceptible) at flowering phase, strengthening the fact that N22 performs better at molecular level also during elevated temperature. This study shows that elevated temperature response is complex and involves multiple biological processes which are needed to be characterized to address the challenges of future climate extreme conditions

Whole-Genome Sequencing of KMR3 and Oryza rufipogon-Derived Introgression Line IL50-13 (Chinsurah Nona 2/Gosaba 6) Identifies Candidate Genes for High Yield and Salinity Tolerance in Rice

The genomes of an elite rice restorer line KMR3 (salinity-sensitive) and its salinity-tolerant introgression line IL50-13, a popular variety of coastal West Bengal, India, were sequenced. High-quality paired-end reads were obtained for KMR3 (147.6 million) and IL50-13 (131.4 million) with a sequencing coverage of 30X-39X. Scaffolds generated from the pre-assembled contigs of each sequenced genome were mapped separately onto the reference genome of Oryza sativa ssp. japonica cultivar Nipponbare to identify genomic variants in terms of SNPs and InDels. The SNPs and InDels identified for KMR3 and IL50-13 were then compared with each other to identify polymorphic SNPs and InDels unique and common to both the genomes. Functional enrichment analysis of the protein-coding genes with unique InDels identified GO terms involved in protein modification, ubiquitination, deubiquitination, peroxidase activity, and antioxidant activity in IL50-13. Linoleic acid metabolism, circadian rhythm, and alpha-linolenic acid metabolism pathways were enriched in IL50-13. These GO terms and pathways are involved in reducing oxidative damage, thus suggesting their role in stress responses. Sequence analysis of QTL markers or genes known to be associated with grain yield and salinity tolerance showed polymorphism in 20 genes, out of which nine were not previously reported. These candidate genes encoded Nucleotide-binding adaptor shared by APAF-1, R proteins, and CED-4 (NB-ARC) domain-containing protein, cyclase, receptor-like kinase, topoisomerase II-associated protein PAT1 domain-containing protein, ion channel regulatory protein, UNC-93 domain-containing protein, subunit A of the heteromeric ATP-citrate lyase, and three conserved hypothetical genes. Polymorphism was observed in the coding, intron, and untranslated regions of the genes on chromosomes 1, 2, 4, 7, 11, and 12. Genes showing polymorphism between the two genomes were considered as sequence-based new candidates derived from Oryza rufipogon for conferring high yield and salinity tolerance in IL50-13 for further functional studies

Genetic characterization and population structure of Indian rice cultivars and wild genotypes using core set markers

Genetic diversity among 23 rice genotypes including wild species and cultivars of indica, japonica, aus and aromatic type was investigated using 165 genomewide core set microsatellite (SSR) markers. This genotypic characterization was undertaken to know the genetic similarity among the parental lines to be used in developing a set of chromosome segment substitution lines. In all, 253 alleles were identified using 77 polymorphic SSRs, and polymorphism information content ranged from 0.31 to 0.97 with a mean of 0.79. Cluster analysis grouped the genotypes into three clusters at a genetic similarity of 0.26–0.75. Wild accessions grouped together in cluster-I, indica cultivars formed cluster-II, and aromatic, japonica and aus types came under cluster-III. Principal component analysis also showed similar results. The genotypic data was analyzed using STRUCTURE, and genotypes were grouped into four populations. RM1018 on chromosome 4, RM8009 on chromosome 7, and RM273 on chromosome 12 amplified alleles specific to wild accessions. The information obtained from core set markers would help in selecting diverse parents including wild accessions and for tracking alleles in mapping or breeding populations. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s13205-016-0409-7) contains supplementary material, which is available to authorized users

Increased Catalase Activity and Maintenance of Photosystem II Distinguishes High-Yield Mutants From Low-Yield Mutants of Rice var. Nagina22 Under Low-Phosphorus Stress

An upland rice variety, Nagina22 (N22) and its 137 ethyl methanesulfonate (EMS)-induced mutants, along with a sensitive variety, Jaya, was screened both in low phosphorus (P) field (Olsen P 1.8) and in normal field (Olsen P 24) during dry season. Based on the grain yield (YLD) of plants in normal field and plants in low P field, 27 gain of function (high-YLD represented as hy) and 9 loss of function (low-YLD represented as ly) mutants were selected and compared with N22 for physiological and genotyping studies. In low P field, hy mutants showed higher P concentration in roots, leaves, grains, and in the whole plant than in ly mutants at harvest. In low P conditions, Fv/Fm and qN were 24% higher in hy mutants than in ly mutants. In comparison with ly mutants, the superoxide dismutase (SOD) activity in the roots and leaves of hy mutants in low P fields was 9% and 41% higher at the vegetative stage, respectively, but 51% and 14% lower in the roots and leaves at the reproductive stage, respectively. However, in comparison with ly mutants, the catalase (CAT) activity in the roots and leaves of hy mutants in low P fields was 35% higher at the vegetative stage and 15% and 17% higher at the reproductive stage, respectively. Similarly, hy mutants in low P field showed 20% and 80% higher peroxidase (POD) activity in the roots and leaves at the vegetative stage, respectively, but showed 14% and 16% lower POD activity at the reproductive stage in the roots and leaves, respectively. Marker trait association analysis using 48 simple sequence repeat (SSR) markers and 10 Pup1 gene markers showed that RM3648 and RM451 in chromosome 4 were significantly associated with grain YLD, tiller number (TN), SOD, and POD activities in both the roots and leaves in low P conditions only. Similarly, RM3334 and RM6300 in chromosome 5 were associated with CAT activity in leaves in low P conditions. Notably, grain YLD was positively and significantly correlated with CAT activity in the roots and shoots, Fv/Fm and qN in low P conditions, and the shoots’ P concentration and qN in normal conditions. Furthermore, CAT activity in shoots was positively and significantly correlated with TN in both low P and normal conditions. Thus, chromosomal regions and physiological traits that have a role in imparting tolerance to low P in the field were identified

Characterization of a Nagina22 rice mutant for heat tolerance and mapping of yield traits

Abstract Background Heat is one of the major factors that considerably limit rice production. Nagina 22 (N22) is a deep-rooted, drought and heat tolerant aus rice cultivar. This study reports the characterization of a previously isolated dark green leaf mutant N22-H-dgl219 (NH219) which showed reduced accumulation of reactive oxygen species in leaf under 40°C heat conditions. The mutant was characterized for several traits in field under ambient (38°C) and heat stress (44°C) conditions by raising temperature artificially from flowering stage till maturity by covering plants with polythene sheets during dry season 2011. Yield traits were mapped in 70 F2 segregants of IR64 × NH219 and 36 F2 segregants of its reciprocal cross. Results Leaf proteome analysis using two-dimensional gel electrophoresis from N22 and NH219 showed distinct constitutive expression of ribulose bisphosphate carboxylase large chain precursor (EC 4.1.1.39) in NH219 under ambient growth condition. Heat stress resulted in reduction of all 11 traits except plant height in both N22 and NH219. The extent of reduction was more in N22 than in NH219. Both pollen viability and spikelet fertility were not reduced significantly in N22 and NH219 but reduced by 20% in IR64. Conclusion NH219 is more tolerant to heat stress than wild type N22 as its percent yield reduction is lesser than N22. Single marker analysis showed significant association of RM1089 with number of tillers and yield per plant, RM423 with leaf senescence, RM584 with leaf width and RM229 with yield per plant. </jats:sec

Phenotyping and Genotype × Environment Interaction of Resistance to Leaffolder, Cnaphalocrocis medinalis Guenee (Lepidoptera: Pyralidae) in Rice

Rice leaffolder, Cnaphalocrocis medinalis is one of the key foliage feeding insects of great concern throughout Asia as it results in significant yield losses. High visibility of damage is triggering farmers to apply toxic pesticides for its management. Therefore, it is vital to identify new stable sources of resistance for leaffolder. Phenotyping of 160 recombinant inbred lines (RILs) of a cross between a resistant parent, W1263 and a susceptible parent, TN1 using a rapid field screening method for three seasons resulted in identification of nine RILs as stable sources of resistance to rice leaffolder. Phenotypic frequency distributions were found continuous indicating that the resistance is a quantitative trait governed by polygenes. Phenotypic data for three seasons were analyzed using Genotype and Genotype × Environment Interaction (GGE) analysis for identification of stable resistant lines. Additive main effect and multiplicative interaction (AMMI) analysis showed that 86.41% of the total sum of square of damaged leaf area was attributed to genotype (GEN) effect; 0.48% to environment (ENV) effects and 5.68% to genotype by environment (G × E) interaction effects. Damage area, damage score and leaf length showed very high broad-sense heritability across three environments. However, leaf width had low heritability indicating higher environment influence. Phylogenetic analysis grouped these 160 RILs and parents into five clusters based on resistant reaction. AMMI and GGE biplot analysis revealed that stable genotypes G8 (MP114) and G3 (MP108) with lower damage area and damage score can be utilized in developing cultivars with leaffolder resistance

Whole Genome Characterization of a Few EMS-Induced Mutants of Upland Rice Variety Nagina 22 Reveals a Staggeringly High Frequency of SNPs Which Show High Phenotypic Plasticity Towards the Wild-Type

The Indian initiative, in creating mutant resources for the functional genomics in rice, has been instrumental in the development of 87,000 ethylmethanesulfonate (EMS)-induced mutants, of which 7,000 are in advanced generations. The mutants have been created in the background of Nagina 22, a popular drought- and heat-tolerant upland cultivar. As it is a pregreen revolution cultivar, as many as 573 dwarf mutants identified from this resource could be useful as an alternate source of dwarfing. A total of 541 mutants, including the macromutants and the trait-specific ones, obtained after appropriate screening, are being maintained in the mutant garden. Here, we report on the detailed characterizations of the 541 mutants based on the distinctness, uniformity, and stability (DUS) descriptors at two different locations. About 90% of the mutants were found to be similar to the wild type (WT) with high similarity index (&gt;0.6) at both the locations. All 541 mutants were characterized for chlorophyll and epicuticular wax contents, while a subset of 84 mutants were characterized for their ionomes, namely, phosphorous, silicon, and chloride contents. Genotyping of these mutants with 54 genomewide simple sequence repeat (SSR) markers revealed 93% of the mutants to be either completely identical to WT or nearly identical with just one polymorphic locus. Whole genome resequencing (WGS) of four mutants, which have minimal differences in the SSR fingerprint pattern and DUS characters from the WT, revealed a staggeringly high number of single nucleotide polymorphisms (SNPs) on an average (16,453 per mutant) in the genic sequences. Of these, nearly 50% of the SNPs led to non-synonymous codons, while 30% resulted in synonymous codons. The number of insertions and deletions (InDels) varied from 898 to 2,595, with more than 80% of them being 1–2 bp long. Such a high number of SNPs could pose a serious challenge in identifying gene(s) governing the mutant phenotype by next generation sequencing-based mapping approaches such as Mutmap. From the WGS data of the WT and the mutants, we developed a genic resource of the WT with a novel analysis pipeline. The entire information about this resource along with the panicle architecture of the 493 mutants is made available in a mutant database EMSgardeN22 (http://14.139.229.201/EMSgardeN22)