Claudio Brondani

Grain yield is a highly complex quantitative trait in rice, resulting from the interaction of multiple genetic, physiological and environmental factors. Although genome-wide association studies (GWAS) have successfully identified loci associated with grain yield, translating statistical associations into biologically meaningful candidate variants remains a major challenge, particularly for variants located in regulatory regions. This study aimed to identify genomic variants associated with grain yield in upland rice and to develop an integrative framework for functionally prioritizing candidate variants through the combination of genome-wide association analysis, functional annotation and regulatory genomics. A panel of 252 accessions from the Brazilian Rice Core Collection was phenotyped for grain yield and genotyped with 35,763 single nucleotide polymorphism (SNP) markers. GWAS identified 29 SNPs significantly associated with grain yield, including 16 variants located within or near annotated genes and 13 located in intergenic regions. The identified candidate genes were involved in signal perception, metabolite transport, amino acid and energy metabolism, hormone biosynthesis, protein turnover, RNA processing and disease resistance, highlighting the polygenic architecture of grain yield. Functional characterization of the intergenic regions revealed enrichment of cis -regulatory elements recognized by transcription factors associated with hormonal signaling, drought response, carbon metabolism, photosynthesis and reproductive development, indicating that regulatory variation represents an important component of grain yield determination. By integrating GWAS signals, candidate gene annotation, cis -regulatory element characterization and the physical proximity between SNPs and cis -regulatory elements, an integrative prioritization strategy identified seven intergenic SNPs as the most promising candidates for functional validation. Together, these findings establish a robust framework for discovering, prioritizing and functionally validating regulatory variants, bridging the gap between statistical associations and biological function while providing a rational strategy for translating GWAS discoveries into molecular breeding of complex traits.

The objective of this work was to identify quantitative trait loci (QTLs) associated with the traits grain yield, plant height, and flowering, as well as superior inbred lines resulting from the intersubspecific cross between 'Araguaia' (Oryza sativa subsp. japonica) and 'Maninjau' (Oryza sativa subsp. indica) rice population. A population consisting of 234 recombinant inbred lines (RILs) was assessed in two environments and genotyped using single nucleotide polymorphisms (SNPs) and SilicoDArT markers. Twenty-two QTLs accounting for phenotypic variation ranging from 3.94% to 35.36%, were identified as significant, as follows: six for grain yield, five for flowering, and eleven for plant height. New QTLs were consistently identified for height and flowering traits with the SNP marker 12 22887040, in both environments, and highlighted for assisted selection of early rice varieties. In both environments, the RIL 1572 with the greatest productivity (6,581 kg ha⁻¹), precocity of 70 days to flowering, and the lowest plant height (90 cm) is highly recommended for integration into crosses with elite materials from the rice breeding program.

Water deficit significantly affects global crop growth and productivity, particularly in water-limited environments, such as upland rice cultivation, reducing grain yield. Plants activate various defense mechanisms during water deficit, involving numerous genes and complex metabolic pathways. Exploring homologous genes that are linked to enhanced drought tolerance through the use of genomic data from model organisms can aid in the functional validation of target species. We evaluated the upland rice OsCPK5 gene, an A. thaliana AtCPK6 homolog, by overexpressing it in the BRSMG Curinga cultivar. Transformants were assessed using a semi-automated phenotyping platform under two irrigation conditions: regular watering, and water deficit applied 79 days after seeding, lasting 14 days, followed by irrigation at 80% field capacity. The physiological data and leaf samples were collected at reproductive stages R3, R6, and R8. The genetically modified (GM) plants consistently exhibited higher OsCPK5 gene expression levels across stages, peaking during grain filling, and displayed reduced stomatal conductance and photosynthetic rate and increased water-use efficiency compared to non-GM (NGM) plants under drought. The GM plants also exhibited a higher filled grain percentage under both irrigation conditions. Their drought susceptibility index was 0.9 times lower than that of NGM plants, and they maintained a higher chlorophyll a/b index, indicating sustained photosynthesis. The NGM plants under water deficit exhibited more leaf senescence, while the OsCPK5-overexpressing plants retained their green leaves. Overall, OsCPK5 overexpression induced diverse drought tolerance mechanisms, indicating the potential for future development of more drought-tolerant rice cultivars.

Water deficit greatly affects global crop growth and productivity, particularly in water-limited environments like upland rice cultivation, leading to reduced grain yield. Plants activate various defense mechanisms during water deficit, involving numerous genes and complex metabolic pathways. Exploring orthologous genes, linked to enhanced drought tolerance, can aid functional validation in target species using genomic data from model organisms. Here, we studied the OsCPK5 gene in upland rice, an AtCPK6 ortholog from A. thaliana, by overexpressing it in the BRSMG Curinga cultivar. Transformants were assessed under two conditions: water deficit applied 79 days after seeding, lasting 14 days, followed by 7 days of irrigation at 80% field capacity. Physiological data and samples were collected at R3, R6, and R8 stages. GM plants consistently exhibited higher OsCPK5 gene expression across stages, peaking during grain filling. During this phase, GM plants displayed reduced stomatal conductance, photosynthetic rate, and increased water use efficiency compared to non-GM plants under drought. GM plants also exhibited higher filled grain percentage in both irrigation conditions. Their drought susceptibility index was 0.9 times lower than NGM plants, and they maintained a higher chlorophyll a/b index, indicating sustained photosynthesis. NGM plants under water deficit exhibited more leaf senescence, while OsCPK5-overexpressing plants retained green leaves. Overall, OsCPK5 overexpression induced diverse drought tolerance mechanisms, hinting at potential for future development of more drought-tolerant rice cultivars.

The objective of this work was to evaluate the physiological, phenotypic, and gene expression parameters in genetically modified (GM) rice plants that overexpress the Oryza sativa Vacuolar H⁺-Pyrophosphatase 1 (OVP1) gene, compared with non-genetically modified (NGM) rice. GM and NGM plants of the BRSMG Curinga cultivar were evaluated in two experiments, in a laboratory and greenhouse, in a randomized complete block design, with four replicates. Agronomic traits of interest were estimated, and transcriptome analysis and gene expression quantification were carried out. GM plants showed a 31 and 21% higher number of spikelets per panicle and total number of grains per panicle, respectively, in comparison with NGM plants. Physiological changes occurred during the grain-filling stage, in which GM plants presented a photosynthetic rate and carboxylation efficiency 61 and 89% higher than those of NGM plants, respectively. The overexpression of the OVP1 gene favors the upregulation of some photosynthesis genes and the increase in the number of spikelets and in the photosynthetic rate, but does not favor the increase in grain yield.

The objective of this work was to determine whether the overexpression of the nuclear Rubisco large subunit (rbcL) improves the drought tolerance of the genetically modified (GM) BRSMG Curinga upland rice (Oryza sativa) cultivar. GM and non-genetically modified (NGM) plants of the same cultivar were compared under the two following water treatments: well watered (WW) and water deficit (WD). The performance of the agronomic traits of GM plants, including grain yield, was superior to that of NGM plants in both treatments. By quantitative polymerase chain reaction, GM plants show a significantly higher expression of the rbcL gene in both WW and WD, as well as a larger amount of abscisic acid. With the RNAseq analysis, almost three times more upregulated genes are identified in GM plants in stage 2 after water restriction, indicating a greater protection against water deficit. The higher expression of genes related to the protection of the cellular metabolism and a series of physiological alterations may be involved in the increase in the drought tolerance of GM rice plants overexpressing the rbcL gene.

The objective of this work was to identify the quantitative trait loci (QTLs) associated with grain yield in a rice segregant population (GYP). A population of 245 inbred recombinant rice lines from the 'Epagri 108' (Oryza sativa subsp. indica) x 'IRAT 122' (O. sativa subsp. japonica) cross was evaluated at different locations and years and genotyped by single nucletide polymorphism (SNP) markers. A map of 1,592.8 cM was obtained from 9,831 SNPs, identifying 25 QTLs. The following nine SNPs showed stability between the different environments: M1.37719614 and M6.9563117 for GYP; M4.29340056, M5.25588710, M7.29115624, and M12.4534450 for 100-grain weight (HGW); and M1.38398157, M4.28368337, and M7.25991230 for plant height (PH). Six SNPs were not present in the linkage blocks: M6.9563117 and M4.1077080 for GYP; M5.25588710 and M6.8886398 for HGW; and M2.34471005 and M8.5955948 for PH. The M6.9563117 and M5.25588710 SNPs were considered environmentally stable and were not present in the linkage blocks, showing their high potential for use in marker-assisted selection for grain yield in Brazilian rice breeding programs.

Upland rice is cultivated mostly in Latin America and Africa by small farmers and in areas with risk of dry spells. This study evaluated morphophysiological mechanisms of upland rice associated to drought adaptation. A set of 25 upland rice genotypes were grown in a plant phenotyping platform during 2015 and 2017 under regular irrigation and water restriction. We evaluated morphophysiological traits in shoots (vegetative structures growth, gas exchange, water use efficiency, carboxylation efficiency, water status) and roots (length, surface area, volume and diameter), as well as agronomic traits (grain yield and its components). There was a reduction in grain yield by up to 54 % and 58 % in 2015 and 2017, respectively, under water deficit. Five upland rice genotypes with the best yield performances in both water treatments applied were recommended to the upland rice-breeding program: Bico Ganga, BRS Esmeralda, BRSMG Curinga, Guarani, and Rabo de Burro. In this study, morphophysiological traits associated to drought tolerance concerned the plant high capacity to save water in the leaves, low leaf water potential, high ability to reduce vegetative structures, high water use efficiency, high photosynthetic capacity, and improved capacity to absorb water from drying soil, either by osmotic adjustment or additional investment into the root system. Therefore, we concluded that different secondary traits contributed to drought tolerance and should be evaluated along with grain yield to improve efficiency of breeding selection.

The objective of this work was to identify and validate single-nucleotide polymorphism (SNP) markers related to grain yield in rice (Oryza sativa) core collection. The genome-wide association studies (GWAS) methodology was applied for genotyping of 541 rice accessions by 167,470 SNPs. The grain yield of these accessions was estimated through the joint analysis of nine field experiments carried out in six Brazilian states. Fifteen SNPs were significantly associated with grain yield, and out of the ten SNPs converted to TaqMan assays, four discriminated the most productive accessions. These markers were used for the screening of rice accessions with favorable alleles. The selected accessions were, then, evaluated in field experiments in target environments, in order to select the most productive ones. This screening reduces the number of accessions evaluated experimentally, making it possible to prioritize those with higher productive potential, which allows of the increase of the number of replicates and, consequently, of the experimental accuracy.