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Plant breeding efforts to boost rice productivity have focused on developing a haploid development pipeline. CENH3 gene has emerged as a leading player that can be manipulated to engineer haploid induction system. Currently, allele mining for the OsCENH3 gene was done by PCR-based resequencing of 33 wild species accessions of genus Oryza and in silico mining of alleles from pre-existing data. We have identified and characterized CENH3 variants in genus Oryza. Our results indicated that the majority CENH3 alleles present in the Oryza gene pool carry synonymous substitutions. A few non-synonymous substitutions occur in the N-terminal Tail domain (NTT). SNP A/G at position 69 was found in accessions of AA genome and non-AA genome species. Phylogenetic analysis revealed that non-synonymous substitutions carrying alleles follow pre-determined evolutionary patterns. O. longistaminata accessions carry SNPs in four codons along with indels in introns 3 and 6. Fifteen haplotypes were mined from our panel; representative mutant alleles exhibited structural variations upon modeling. Structural analysis indicated that more than one structural variant may be exhibited by different accessions of single species (Oryza barthii). NTT allelic mutants, though not directly implicated in HI, may show variable interactions. HI and interactive behavior could be ascertained in future investigations.

Rice false smut (RFS), an emerging major fungal disease worldwide caused by Ustilaginoidea virens, affects rice grain quality and yield. RFS cause 2.8-49% global yield loss depending upon disease severity and cultivars. In India, the yield loss due to RFS ranged from 2 to 75%. Identification of the genes or quantitative trait loci (QTLs) governing disease resistance would be of utmost importance towards mitigating the economic losses incurred due to RFS. Here, we report mapping of RFS resistance QTLs from a resistant breeding line RYT2668. The mapping population was evaluated for RFS resistance under the field condition in three cropping seasons 2013, 2015, and 2016. A positive correlation among infected panicle/plant, total smut ball/panicle, and disease score was observed in the years 2013, 2015, and the mean data. A total of seven QTLs were mapped on rice chromosomes 2, 4, 5, 7, and 9 using 2326 single nucleotide polymorphism markers. Of these, two QTLs, qRFSr5.3 and qRFSr7.1a, were associated with the infected panicle per plant, one QTL qRFsr9.1 with total smut ball per panicle, and four QTLs qRFSr2.2, qRFSr4.3, qRFSr5.4, and qRFSr7.1b with disease score. Among them, a novel QTL qRFSr9.1 on chromosome 9 exhibits the largest phenotypic effect. The prediction of putative candidate genes within the qRFSr9.1 revealed four nucleotide-binding sites-leucine-rich repeat (NBS-LRR) domain-containing disease resistance proteins. In summary, our findings mark the hotspot region of rice chromosomes carrying genes/QTLs for resistance to the RFS disease.

Sheath blight (ShB) caused by Rhizoctonia solani is one of the serious constraints that hamper global rice production. It causes severe yield loss of up to 69% under favorable environmental conditions. The application of chemical fungicides remains the primary approach for the management of this disease. But, overuse of chemical fungicides causes potential health and environmental risk. The use of host plant resistance is a very effective, economic, and sustainable strategy to control sheath blight disease in rice. In this report, we have evaluated 405 accessions of Oryza rufipogon for four ShB-related traits i.e. lesion height (LH), plant height (PH), relative lesion length (RLH), and disease score (DS) during the years 2015, 2016, and 2017. A total of 44,109 high-quality SNP markers on 301 accessions were employed to identify significant marker-trait associations (MTAs) using a genome-wide association study (GWAS). The GWAS analysis revealed a total of 22 significant SNPs associated with the ShB-related traits distributed on all the rice chromosomes except 10 and 12. Among them, eleven were associated with RLH, seven with PH, and one each with LH and DS. The SNPs on chromosomes 3 and 9 were associated with multiple traits. Furthermore, seven ShB resistance loci were also found to be co-localized with previously reported ShB resistance genes/QTLs. These findings provide valuable information to identify key SNPs associated with ShB resistance. Significantly associated SNPs could be used for introgression of ShB resistance traits into rice cultivars using marker-assisted selection (MAS).

Background Rice (Oryza sativa L.) is the major source of daily caloric intake for more than 30% of the human population. However, the sustained productivity of this staple food crop is continuously threatened by various pathogens and herbivores. Breeding has been successful in utilizing various mechanisms of defense by gene pyramiding in elite cultivars, but the continuous resurgence of highly resistant races of pathogens and herbivores often overcomes the inherent capacity of host plant immunity. MicroRNAs (miRNAs) are endogenous, short, single-stranded, non-coding RNA molecules that regulate gene expression by sequence-specific cleavage of target mRNA or suppressing target mRNA translation. While miRNAs function as upstream regulators of plant growth, development, and host immunity, their direct effects on growth and development in the context of balancing defenses with agronomic potential have not been extensively discussed and explored as a more viable strategy in breeding for disease and pest resistant cultivars of rice with optimal agronomic potentials. Results Using the available knowledge in rice and other model plants, this review examines the important roles of miRNAs in regulating host responses to various fungal, bacterial, and viral pathogens, and insect pests, in the context of gains and trade-offs to crop yield. Gains from R-gene-mediated resistance deployed in modern rice cultivars are often undermined by the rapid breakdown of resistance, negative pleiotropic effects, and linkage drags with undesirable traits. In stark contrast, several classes of miRNAs are known to efficiently balance the positive gains from host immunity without significant costs in terms of losses in agronomic potentials (i.e., yield penalty) in rice. Defense-related miRNAs such as Osa-miR156, Osa-miR159, Osa-miR162, Osa-miR396, Osa-530, Osa-miR1432, Osa-miR1871, and Osa-miR1873 are critical in fine-tuning and integrating immune responses with physiological processes that are necessary to the maintenance of grain yield. Recent research has shown that many defense-related miRNAs regulate complex and agronomically important traits. Conclusions Identification of novel immune-responsive miRNAs that orchestrate physiological processes critical to the full expression of agronomic potential will facilitate the stacking of optimal combinations of miRNA-encoding genes to develop high-yielding cultivars with durable resistance to disease and insect pests with minimal penalties to yield.

Wild relatives of rice (Oryza sativa L.) represent enormous genetic diversity for many traits. Many domestication related traits found only in wild species that are lost from cultivated species during domestication. In this study, we report high-resolution mapping of two domestications related quantitative trait locus (QTL) controlling purple sheath color and plant height using high-density SNP based linkage map in an interspecific F2 population derived from a cross between O. nivara Sharma & Shastry and O. sativa. The segregation ratio of purple sheath color fit into 3:1 indicated that the presence of a dominant gene. The segregation of plant height trait exhibited a normal distribution in F2 and F2:3 generations. QTL analysis showed the identification of two QTLs controlling purple sheath color (designated as qPsh6.1 and qPsh6.2) on chromosome 6 explaining 27.12% and 14% phenotypic variance, respectively. The only plant height QTL (tentatively designated as qPh1.9) was mapped on chromosome 1 contributing 18.3% phenotypic variance. The qPsh6.1 covers 25.17 Kb region flanked by AX-95928721 and AX-95954573 containing five open reading frames (ORFs) while qPsh6.2 spanned in 118.2 Kb region containing 19 ORFs flanked by markers AX-95956260 and AX-95928295. The qPh1.9 gene controlling plant height was mapped between two SNP markers, AX-95944819 and AX-95964796 co-segregated with the locus LOC_Os01g66690 encoding for ZIP4/SPO22 protein. The detailed genetic study of the location of these genes will be useful for the map-based cloning.

A novel recessive bacterial blight resistance locus designated as a xa-45(t) was identified from Oryza glaberrima accession IRGC 102600B, transferred to O. sativa and mapped to the long arm of chromosome 8 using ddRAD sequencing approach. The identified QTL spans 80 kb region on Nipponbare reference genome IRGSP-1.0 and contains 9 candidate genes. An STS marker developed from the locus LOC_Os08g42410 was found co-segregating with the trait and will be useful for marker-assisted transfer of this recessive resistance gene in breeding programs. Bacterial blight, caused by Xanthomonas oryzae pv. oryzae, is one of the major constraints of rice productivity in Southeast Asia. In spite of having 44 bacterial blight resistance genes from cultivated rice and wild species, the durability of resistance is always at stake due to the continually evolving nature of the pathogen and lack of suitable chemical control. Here, we report high-resolution genetic mapping of a novel bacterial blight resistance gene tentatively designated as a xa-45(t) from an introgression line derived from Oryza glaberrima accession IRGC 102600B. This introgression line was crossed with the susceptible rice indica cultivar cv. Pusa 44 to generate F2 and F2:3 populations for inheritance and mapping studies. The inheritance studies revealed the presence of single recessive locus controlling resistance to the Xanthomonas pathotype seven. A high-density linkage map was constructed using double-digest restriction-associated DNA sequencing of 96 F2 populations along with the parents. The QTL mapping identified a major locus on the long arm of rice chromosome 8 with a LOD score of 33.22 between the SNP markers C8.26737175 and C8.26818765. The peak marker, C8.26810477, explains 49.8% of the total phenotypic variance and was positioned at 202.90 cM on the linkage map. This major locus spans 80 kb region on Nipponbare reference genome IRGSP-1.0 and contains 9 candidate genes. A co-segregating STS marker was developed from the LOC_Os08g42410 for efficient transfer of this novel gene to elite cultivars.

Rice bran oil is good quality edible oil, rich in antioxidants and comprised typically of oleic-linoleic type fatty acids. However, presence of a highly lipolytic enzyme Phospholipase D alpha1 (OsPLDα1) increases free fatty acid content in the oil which further leads to stale flavor and rancidity of the oil, making it unfit for human consumption. In this study, we compared the upstream regions of OsPLDα1 orthologs across 34 accessions representing 5 wild Oryza species and 8 cultivars, to uncover sequence variations and identify cis-elements involved in differential transcription of orthologs. Alignment of the upstream sequences to the Nipponbare OsPLDα1 reference sequence revealed the presence of 39 SNPs. Phylogenetic analysis showed that all the selected cultivars and wild species accessions are closely related to the reference except for three accessions of O. rufipogon (IRGC89224, IRGC104425, and IRGC105902). Furthermore, using exon-specific qRT-PCR, OsPLDα1 expression patterns in immature grains indicated significant differences in transcript abundance between the wild species accessions. In comparison to the control, lowest gene expression was observed in IRGC89224 accession (0.20-fold) followed by IRGC105902 (0.26-fold) and IRGC104425 (0.41-fold) accessions. In-silico analysis of the OsPLDα1 promoter revealed that the copy number variations of CGCGBOXAT, GT1CONSENSUS, IBOXCORE, NODCON2GM, OSE2ROOTNODULE, SURECOREATSULTR11, and SORLIP1AT cis-elements play an important role in the transcriptional activities of orthologous genes. Owing to the presence of ARFAT and SEBF elements only in the IRGC89224 accession, which had the lowest gene expression as well, these putative upstream regulatory sequences have been identified as novel cis-elements which may act as repressors in regulating the OsPLDα1 gene expression. The accessions identified with low OsPLDα1 expressions could be further deployed as potential donors of ideal OsPLDα1 allele for transfer of the desired trait into elite rice cultivars.

A BPH-resistant locus designated as Bph34 identified in Oryza nivara acc. IRGC104646 on long arm of chromosome 4 using high-resolution mapping with 50 K SNP chip. BPH resistance contributed by locus showed dominant inheritance in F2 and F3. The Bph34 locus is 91 kb in size and contains 11 candidate genes. In addition to SNP markers, SSR markers, RM16994 and RM17007 co-segregated with the BPH resistance. These two SSR markers can facilitate marker-assisted transfer of the Bph34 locus into elite rice cultivars in all labs. Brown planthopper (BPH, Nilaparvata lugen Stal) is one of the most destructive insects of rice (Oryza sativa L.) causing significant yield losses annually. Exploiting host plant resistance to BPH and incorporating resistant genes in susceptible commercial cultivars is economical and environmentally friendly approach to manage this pest. Here, we report high-resolution mapping of a novel genetic locus for resistance to BPH, designated as Bph34 on long arm of rice chromosome 4. The locus was mapped using an interspecific F2 population derived from a cross between susceptible indica cultivar PR122 and BPH-resistant wild species, O. nivara acc. IRGC104646. Inheritance studies performed using F2 and F2:3 populations revealed the presence of single dominant gene. Construction of high-density linkage map using 50 K SNP chip (OsSNPnks) followed by QTL mapping identified single major locus at 28.8 LOD score between SNP markers, AX-95952039 and AX-95921548. The major locus contributing resistance to BPH designated as Bph34 and explained 68.3% of total phenotypic variance. The Bph34 locus is 91 Kb in size on Nipponbare reference genome-IRGSP-1.0 and contains 11 candidate genes. In addition to associated SNP markers, two SSR markers, RM16994 and RM17007, also co-segregated with the Bph34 which can be used efficiently for markers assisted transfer into elite rice cultivars across the labs.

We evaluated automated OCT-derived drusen volume measures in a population-based study (n = 4,512) aged ≥40 years, and its correlation with conventional color fundus photographs (CFP)-derived early AMD features. Participants had protocol-based assessment to capture medical and ocular history, genotyping for SNPs in CFH, ARMS2, and CETP, CFP-based AMD grading and automated drusen volume based on SD-OCT using built-in software (Cirrus OCT advanced RPE analysis software). Significantly fewer eyes with early AMD features (drusen, hyperpigmentation, soft or reticular drusen) had drusen volume = 0 mm3 (p 0 mm3, increasing AMD severity was associated with increase in drusen volume (correlation coefficient 0.17, p 3. Factors associated with drusen volume included age (OR 1.42 per 5 years, 95% confidence interval [CI] 2.76, 4.48), systolic blood pressure (OR1.00, 95% CI 1.00, 1.01), ethnic Malay (OR 1.54, 95% CI 1.29, 1.83) and Chinese (OR 1.66, 95% CI 1.37, 2.01) compared to Indian. The ARMS2 rs10490924 T allele was associated with increased drusen volume in subjects with AMD (multivariable adjusted OR1.54, 95% CI 1.08, 2.19). Automated OCT-derived drusen volume is correlated with CFP-based AMD grading in many, but not all subjects. However the agreement is not good. These two modalities provide complementary information and should be incorporated into future studies.

Increasing the rice productivity from the current 10 to 12 tons/ha to meet the demand of estimated 8.8 billion people in 2035 is posing a major challenge. Wild relatives of rice contain some novel genes which can help in improving rice yield. Spikelet per panicle (SPP) is a valuable trait for determining yield potential in rice. In this study, a major QTL for increasing SPP has been identified, mapped, and transferred from African wild rice O. longistaminata to O. sativa (L.). The QTL was mapped on the long arm of chromosome 2 in a 167.1 kb region flanked by SSR markers RM13743 and RM13750, which are 1.0 cM apart, and is designated as qSPP2.2. The QTL explained up to 30% of phenotypic variance in different generations/seasons and showed positive additive effect of allele contributed by O. longistaminata. In addition, O. longistaminata allele in qSPP2.2 contributed to increase in grains per panicle, but decrease in the tillers per plant. The 167.1 kb region contains 23 predicted genes. Based on the functional annotation, three genes, LOC_Os02g44860, LOC_Os02g44990, and LOC_Os02g45010, were selected as putative candidates for characterization. Sequence analysis of the three genes revealed functional variations between the parental lines for LOC_Os02g44990 and a variation in 5′UTR for LOC_Os02g45010 which will help further to identify putative candidate gene(s). This is the first yield component QTL to be identified, mapped, and transferred from O. longistaminata.

We aimed to determine the 6-year incidence and risk factors of age-related macular degeneration (AMD) in first and second generations of Singaporean Indians. Baseline examination was conducted in 2007–9 and 6-year propsective follow-up examination of this Indian population in 2013–5. All participants underwent interviews with questionnaires and comprehensive medical and eye examinations. Incidence was age-standardized to Singaporean 2010 census. Risk factors associated with AMD incidence were assessed and compared between first and second generations of immigrants. Among 2200 persons who participated in the follow-up examination (75.5% response rate), gradable fundus photographs were available in 2105. The 6-year age-standardized incidences of early and late AMD were 5.26% and 0.51% respectively. Incident early AMD was associated with cardiovascular disease history (HR 1.59, 95% CI 1.04–2.45), underweight body mass index (BMI) (HR 3.12, 95% CI 1.37–7.14) (BMI of

Development and utilization of genetic markers play a pivotal role in marker-assisted breeding of wheat cultivars with pyramids of disease resistance genes. The objective of this study was to develop a closed-tube, gel-free assay for high-throughput genotyping of leaf rust resistance locus Lr21. Polymorphism identified from re-sequencing of a 2.4-kb fragment covering the functional region of the Lr21 gene from the second to the fourth indels was targeted for assay development. The generated sequence data revealed the 88- or 105-bp indel in the first intron of the Lr21 gene in the selected resistant cultivars compared to susceptible US spring and winter wheat cultivars. Allele-specific primers for a KASPar assay were designed around the junction of the indel at position 1,346 bp. The marker was tested on a panel of 384 US wheat lines and found to be effective in differentiating resistant and susceptible genotypes.

To quantify the fundus color on digital color fundus photographs as a surrogate of myopic chorioretinal degeneration in myopic subjects using a novel computer software program. In this cross-sectional study, the following details were recorded in 152 eyes of 78 myopic subjects: Snellen visual acuity, refractive status, ocular biometric parameters, intraocular pressure, color fundus photography, and myopic chorioretinal degeneration index (MCDI) using an automated computer program. The mean age of the subjects was 32.58 ± 11.12 years (range, 21-62 years). Subjects with aged ≥40 years had a significantly higher MCDI when compared with subjects with aged

Iron and zinc deficiency affects more than half of the world population due to low inherent micronutrient content of cereals and other staple foods. The micronutrient deficiency is further aggravated by poor availability of these minerals in calcareous soils and their uptake by crop plants. Series of available wheat-Aegilops addition lines were evaluated for identification of alien chromosomes carrying genes for high grain iron and zinc concentrations and release of mugineic acid(s) facilitating micronutrient uptake under their deficient conditions. Addition lines of chromosome 2Sv, 2Uv and 7Uv of Ae. peregrina, 2Sl and 7Sl of Ae. longissima and 2U of Ae. umbellulata were found to carry genes for high grain iron whereas the group 7 chromosomes had genes for higher grain zinc. Higher release of mugineic acid (MA) under iron deficient condition was observed in addition lines of chromosome 2Sv, 2Uv, 4Uv and 7Sv of Ae. peregrina, 2Sl and 6Sl of Ae. longissima and 2U and 5U of Ae. umbellulata. Higher grain and...

Dietary deficiency of iron and zinc micronutrients affects more than two billion people worldwide. Breeding for micronutrient-dense crops is the most sustainable and cost-effective approach for alleviation of micronutrient malnutrition. Three accessions of Aegilops peregrina (Hack.) Maire & Weill (2n = 28, UPUPSPSP), selected for high grain iron and zinc concentration were crossed with Triticum aestivum L. cv. Chinese Spring (Ph I ). The sterile F1 hybrids were backcrossed with elite wheat cultivars to get fertile BC2F2 derivatives. Some of the fertile BC2F2 derivatives showed nearly 100% increase in grain iron and more than 200% increase in grain zinc concentration compared to the recipient wheat cultivars. The development of derivatives with significantly higher grain micronutrients, high thousand-grain weight and harvest index suggests that the enhanced micronutrient concentration is due to the distinct genetic system of Ae. peregrina and not to the concentration effect. Genomic in situ hybridization, comparison of introgressed chromosomes with the standard karyotype of Ae. peregrina and simple sequence repeat marker analysis revealed the introgression of 7SP chromosomes in five selected derivatives, 7UP in four, group 4 and 4SP in three and a translocated 5UP of Ae. peregrina in one of the selected derivatives. Molecular marker analysis using the introgressed chromosome markers indicated that two of the BC2F3 progenies were stabilized as disomic addition lines. It could, therefore, be concluded that the group 4 and 7 chromosomes of Ae. peregrina carry the genes for high grain iron and zinc concentration.

Biofortification through genetic manipulation is the best approach for improving micronutrient content of the staple food crops to alleviate hidden hunger, namely, the deficiency of Fe and Zn affecting more than two billion people worldwide. An interspecific hybridization was made between T. aestivum line Chinese Spring (CS) and Aegilops kotschyi accession 3790 selected for high grain iron and zinc concentration. The CS x Ae. kotschyi F(1) hybrid with low chromosome pairing was highly male and female sterile. This was backcrossed with wheat cultivars to get seed set. The selfed BC(1)F(1) and BC(2)F(1) plants with high grain iron and zinc concentration were selected in subsequent generations. The selected derivatives showed 60-136% enhanced grain iron and zinc concentration and 50-120% increased iron and zinc content per seed as compared to the recipient wheat cultivars. Thirteen cytologically stable, fertile and agronomically superior plants with high grain iron and zinc concentrations were selected for molecular characterization. The application of anchored wheat SSR markers, transferable to Ae. kotschyi, to the high grain iron and zinc containing derivatives indicated introgression of group 2 and group 7 chromosomes of Ae. kotschyi. GISH and FISH analysis of some derivatives confirmed the substitution of chromosomes 2S and 7U for their homoeologues of the A genome, suggesting that some of the genes controlling high grain micronutrient content in the Ae. kotschyi accession are on these chromosomes.