OsLRR-RLP2 Gene Regulates Immunity to Magnaporthe oryzae in Japonica Rice.

Rice is an important cereal crop worldwide, the growth of which is affected by rice blast disease, caused by the fungal pathogen Magnaporthe oryzae. As climate change increases the diversity of pathogens, the disease resistance genes (R genes) in plants must be identified. The major blast-resistance genes have been identified in indica rice varieties; therefore, japonica rice varieties with R genes now need to be identified. Because leucine-rich repeat (LRR) domain proteins possess R-gene properties, we used bioinformatics analysis to identify the rice candidate LRR domain receptor-like proteins (OsLRR-RLPs). OsLRR-RLP2, which contains six LRR domains, showed differences in the DNA sequence, containing 43 single-nucleotide polymorphisms (SNPs) in indica and japonica subpopulations. The results of the M. oryzae inoculation analysis indicated that indica varieties with partial deletion of OsLRR-RLP2 showed susceptibility, whereas japonica varieties with intact OsLRR-RLP2 showed resistance. The oslrr-rlp2 mutant, generated using clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9), showed increased pathogen susceptibility, whereas plants overexpressing this gene showed pathogen resistance. These results indicate that OsLRR-RLP2 confers resistance to rice, and OsLRR-RLP2 may be useful for breeding resistant cultivars.

Proteome profiling highlights mechanisms underlying pigment and tocopherol accumulation in red and black rice seeds.

Rice is a major component of the human diet and feeds more than 50 million people across the globe. We previously developed two pigmented rice cultivars, Super-hongmi (red seeds) and Super-jami (black seeds), that are highly rich in antioxidants and exhibit high levels of radical scavenging activities. However, the molecular mechanism underlying the accumulation of pigments and different antioxidants in these rice cultivars remains largely elusive. Here, we report the proteome profiles of mature Super-hongmi and Super-jami seeds, and compared them with the Hopum (white seeds) using a label-free quantitative proteomics approach. This approach led to the identification of 5127 rice seed proteins of which 1628 showed significant changes in the pigmented rice cultivar(s). The list of significantly modulated proteins included a phytoene desaturase (PDS3) which suggested accumulation of ζ-carotene in red seeds while the black seeds seem to accumulate more of anthocyanins because of the higher abundance of dihydroflavonol 4-reductase. Moreover, proteins associated with lignin and tocopherol biosynthesis were highly increased in both red and black cultivars. Taken together, these data report the seed proteome of three different colored rice seeds and identify novel components associated with pigment accumulation in rice.

Cytokinin increases vegetative growth period by suppressing florigen expression in rice and maize.

Increasing the vegetative growth period of crops can increase biomass and grain yield. In rice (Oryza sativa), the concentration of trans -zeatin, an active cytokinin, was high in the leaves during vegetative growth and decreased rapidly upon induction of florigen expression, suggesting that this hormone is involved in the regulation of the vegetative phase. To elucidate whether exogenous cytokinin application influences the length of the vegetative phase, we applied 6-benzylaminopurine (BAP) to rice plants at various developmental stages. Our treatment delayed flowering time by 8-9 days when compared with mock-treated rice plants, but only at the transition stage when the flowering signals were produced. Our observations also showed that flowering in the paddy field is delayed by thidiazuron, a stable chemical that mimics the effects of cytokinin. The transcript levels of florigen genes Heading date 3a (Hd3a) and Rice Flowering locus T1 (RFT1) were significantly reduced by the treatment, but the expression of Early heading date 1 (Ehd1), a gene found directly upstream of the florigen genes, was not altered. In maize (Zea mays), similarly, BAP treatment increased the vegetative phage by inhibiting the expression of ZCN8, an ortholog of Hd3a. We showed that cytokinin treatment induced the expression of two type-A response regulators (OsRR1 and OsRR2) which interacted with Ehd1, a type-B response regulator. We also observed that cytokinin did not affect flowering time in ehd1 knockout mutants. Our study indicates that cytokinin application increases the duration of the vegetative phase by delaying the expression of florigen genes in rice and maize by inhibiting Ehd1.

Global Identification and Characterization of C2 Domain-Containing Proteins Associated with Abiotic Stress Response in Rice (Oryza sativa L.).

C2 domain-containing proteins (C2DPs) have been identified in different genomes that contain single or multiple C2 domains in their C- or N-terminal. It possesses higher functional activity in the transmembrane regions. The identification of C2 domains were reported in a previous study, such as multiple C2 domains and transmembrane-region proteins (MCTPs) and N-terminal-TM-C2 domain proteins (NTMC2s) of rice, Arabidopsis thaliana, and cotton, whereas the C2DP gene family in rice has not been comprehensively studied, and the role of the C2DP gene in rice in response to abiotic stress is not yet fully understood. In this study, we identified 82 C2DPs in the rice genome and divided them into seven groups through phylogenetic analysis. The synteny analysis revealed that duplication events were either exhibited within the genome of rice or between the genomes of rice and other species. Through the analysis of cis-acting elements in promoters, expression profiles, and qRT-PCR results, the functions of OsC2DPs were found to be widely distributed in diverse tissues and were extensively involved in phytohormones-related and abiotic stresses response in rice. The prediction of the microRNA (miRNA) targets of OsC2DPs revealed the possibility of regulation by consistent miRNAs. Notably, OsC2DP50/51/52 as a co-tandem duplication exhibited similar expression variations and involved the coincident miRNA-regulation pathway. Moreover, the results of the genotypic variation and haplotype analysis revealed that OsC2DP17, OsC2DP29, and OsC2DP49 were associated with cold stress responses. These findings provided comprehensive insights for characterizations of OsC2DPs in rice as well as for their roles for abiotic stress.

A new SNP in cyOsPPDK gene is associated with floury endosperm in Suweon 542.

Pyruvate orthophosphate dikinase (PPDK) is a component of glycolysis to mediate endosperm energy charge by adjusting the ratio of ATP to ADP and AMP that proposed to balance the flow of carbon into starch, protein, fatty acid and amino acid biosynthesis. However, these were inconsistent with the first report of a T-DNA insertional knockout mutant of the rice PPDK gene (flo4) showed that rice with inactivated PPDK gene failed to produce a opaque seeds. Therefore, the PPDK might have multifaceted functions in grain filling stage, which in some ways might depend on the direction of the reversible catalysis. Suweon 542 is a rice (Oryza sativa L.) mutant developed from Oryza sativa ssp. japonica cv. Namil. Suweon 542 has a milky-white floury endosperm suitable for dry filling, with low starch damage, low grain hardness, and fine flour particle size. The mutant locus on chromosome 5 controls the floury endosperm phenotype of Suweon 542. Fine mapping of this locus is required for efficient breeding of rice germplasm suitable for dry milling. In this study, whole genome of Suweon 542 and Milyang 23 were re-sequenced using Illumina HiSeq 2500. Co-segregation analysis of F3:4 family populations derived from Suweon 542/Milyang 23 was performed using eight CAPS markers and phenotypic evaluation of the endosperm. The target region was mapped to a 33 kb region and identified to encode cytosolic pyruvate orthophosphate dikinase protein (cyOsPPDK). A G→A SNP in exon 8 of cyOsPPDK resulting in a missense mutation from Gly to Asp at amino acid position 404 was responsible for the floury endosperm of Suweon 542. qRT-PCR experiments revealed that FLO4-4 was expressed to a considerably higher level in Suweon 542 than in Namil during the grain filling stage. Overall, fine mapping of FLO4-4 and candidate gene analysis provided further insight into the floury endosperm of rice, and reveal a novel SNP in cyOsPPDK gene can affect the floury endosperm phenotype through active PPDK gene during grain filling stage.

Genome-wide resequencing of KRICE_CORE reveals their potential for future breeding, as well as functional and evolutionary studies in the post-genomic era.

BACKGROUND: Rice germplasm collections continue to grow in number and size around the world. Since maintaining and screening such massive resources remains challenging, it is important to establish practical methods to manage them. A core collection, by definition, refers to a subset of the entire population that preserves the majority of genetic diversity, enhancing the efficiency of germplasm utilization. RESULTS: Here, we report whole-genome resequencing of the 137 rice mini core collection or Korean rice core set (KRICE_CORE) that represents 25,604 rice germplasms deposited in the Korean genebank of the Rural Development Administration (RDA). We implemented the Illumina HiSeq 2000 and 2500 platform to produce short reads and then assembled those with 9.8 depths using Nipponbare as a reference. Comparisons of the sequences with the reference genome yielded more than 15 million (M) single nucleotide polymorphisms (SNPs) and 1.3 M INDELs. Phylogenetic and population analyses using 2,046,529 high-quality SNPs successfully assigned rice accessions to the relevant rice subgroups, suggesting that these SNPs capture evolutionary signatures that have accumulated in rice subpopulations. Furthermore, genome-wide association studies (GWAS) for four exemplary agronomic traits in the KRIC_CORE manifest the utility of KRICE_CORE; that is, identifying previously defined genes or novel genetic factors that potentially regulate important phenotypes. CONCLUSION: This study provides strong evidence that the size of KRICE_CORE is small but contains high genetic and functional diversity across the genome. Thus, our resequencing results will be useful for future breeding, as well as functional and evolutionary studies, in the post-genomic era.

Construction of a genetic linkage map and analysis of quantitative trait loci associated with the agronomically important traits of Pleurotus eryngii.

Breeding new strains with improved traits is a long-standing goal of mushroom breeders that can be expedited by marker-assisted selection (MAS). We constructed a genetic linkage map of Pleurotus eryngii based on segregation analysis of markers in postmeiotic monokaryons from KNR2312. In total, 256 loci comprising 226 simple sequence-repeat (SSR) markers, 2 mating-type factors, and 28 insertion/deletion (InDel) markers were mapped. The map consisted of 12 linkage groups (LGs) spanning 1047.8cM, with an average interval length of 4.09cM. Four independent populations (Pd3, Pd8, Pd14, and Pd15) derived from crossing between four monokaryons from KNR2532 as a tester strain and 98 monokaryons from KNR2312 were used to characterize quantitative trait loci (QTL) for nine traits such as yield, quality, cap color, and earliness. Using composite interval mapping (CIM), 71 QTLs explaining between 5.82% and 33.17% of the phenotypic variations were identified. Clusters of more than five QTLs for various traits were identified in three genomic regions, on LGs 1, 7 and 9. Regardless of the population, 6 of the 9 traits studied and 18 of the 71 QTLs found in this study were identified in the largest cluster, LG1, in the range from 65.4 to 110.4cM. The candidate genes for yield encoding transcription factor, signal transduction, mycelial growth and hydrolase are suggested by using manual and computational analysis of genome sequence corresponding to QTL region with the highest likelihood odds (LOD) for yield. The genetic map and the QTLs established in this study will help breeders and geneticists to develop selection markers for agronomically important characteristics of mushrooms and to identify the corresponding genes.

A quantitative shotgun proteomics analysis of germinated rice embryos and coleoptiles under low-temperature conditions.

BACKGROUND: At low temperatures, rice grains have a reduced germination rate and grow more slowly, which delays the emergence of rice seedlings from the paddy water surface and significantly increases seedling mortality. In this study, we conducted a shotgun proteomics analysis of geminated embryos and coleoptiles to compare the proteome expression pattern between the low-temperature resistant variety, Tong 88-7, and the low-temperature susceptible variety, Milyang 23. RESULTS: In a shotgun proteomics analysis of low-temperature resistant and susceptible embryos and coleoptiles in both cold and control temperatures, we discovered a total of 2626 non-redundant proteins, with a 0.01 false discovery rate. A comparison of protein expression patterns between resistant and susceptible embryos and coleoptiles under low-temperature and normal conditions revealed that 85 proteins and 196 proteins were expressed by the resistant and susceptible strains, respectively, in response to low temperature. Among them, 12 proteins overlapped. Proteins involved in stress responses, metabolism, and gene expression were expressed in both strains. CONCLUSIONS: Similar molecular functions of the response were detected, suggesting that the resistant and susceptible strain have a similar proteome response to cold temperatures. The resistance of Tong 88-7 to cold-water germination may result from the efficiency of these proteins, rather than activation of additional or different molecular processes. A comparison of protein expression between the resistant and susceptible strains' responses revealed that the more successful low-temperature germination of Tong 88-7 was associated with gibberellin signaling, protein trafficking, and the ABA-mediated stress response.

Synergizing breeding strategies via combining speed breeding, phenotypic selection, and marker-assisted backcrossing for the introgression of Glu-B1i in wheat.

Wheat is a major food crop that plays a crucial role in the human diet. Various breeding technologies have been developed and refined to meet the increasing global wheat demand. Several studies have suggested breeding strategies that combine generation acceleration systems and molecular breeding methods to maximize breeding efficiency. However, real-world examples demonstrating the effective utilization of these strategies in breeding programs are lacking. In this study, we designed and demonstrated a synergized breeding strategy (SBS) that combines rapid and efficient breeding techniques, including speed breeding, speed vernalization, phenotypic selection, backcrossing, and marker-assisted selection. These breeding techniques were tailored to the specific characteristics of the breeding materials and objectives. Using the SBS approach, from artificial crossing to the initial observed yield trial under field conditions only took 3.5 years, resulting in a 53% reduction in the time required to develop a BC2 near-isogenic line (NIL) and achieving a higher recurrent genome recovery of 91.5% compared to traditional field conditions. We developed a new wheat NIL derived from cv. Jokyoung, a leading cultivar in Korea. Milyang56 exhibited improved protein content, sodium dodecyl sulfate-sedimentation value, and loaf volume compared to Jokyoung, which were attributed to introgression of the Glu-B1i allele from the donor parent, cv. Garnet. SBS represents a flexible breeding model that can be applied by breeders for developing breeding materials and mapping populations, as well as analyzing the environmental effects of specific genes or loci and for trait stacking.

Acceleration of wheat breeding: enhancing efficiency and practical application of the speed breeding system.

BACKGROUND: Crop breeding should be accelerated to address global warming and climate change. Wheat (Triticum aestivum L.) is a major food crop. Speed breeding (SB) and speed vernalization (SV) techniques for spring and winter wheat have recently been established. However, there are few practical examples of these strategies being used economically and efficiently in breeding programs. We aimed to establish and evaluate the performance of a breeder-friendly and energy-saving generation acceleration system by modifying the SV + SB system. RESULTS: In this study, a four-generation advancement system for wheat (regardless of its growth habits) was established and evaluated using an energy-efficient extended photoperiod treatment. A glasshouse with a 22-hour photoperiod that used 10 h of natural sunlight and 12 h of LED lights, and minimized temperature control during the winter season, was successful in accelerating generation. Even with one or two field tests, modified speed breeding (mSB) combined with a speed vernalization system (SV + mSB) reduced breeding time by more than half compared to traditional field-based methods. When compared to the existing SV + SB system, the SV + mSB system reduced energy use by 80% to maintain a 22-hour photoperiod. Significant correlations were found between the SV + mSB and field conditions in the number of days to heading (DTH) and culm length (CL). Genetic resources, recombinant inbred lines, and breeding materials that exhibited shorter DTH and CL values under SV + mSB conditions showed the same pattern in the field. CONCLUSIONS: The results of our SV + mSB model, as well as its practical application in wheat breeding programs, are expected to help breeders worldwide incorporate generation acceleration systems into their conventional breeding programs.

Tumor budding as a predictor of disease-free survival in patients with cholangiocarcinoma.

Background: Tumor budding is considered a prognostic factor in several solid cancer types. However, we lack comprehensive information on the importance of tumor budding in cholangiocarcinoma. Therefore, we aimed to assess the prognostic value of tumor budding in intrahepatic and extrahepatic cholangiocarcinomas and to evaluate its correlations with other clinicopathological parameters. Methods: We monitored 219 patients who underwent surgery for intrahepatic or extrahepatic cholangiocarcinoma at the Pusan National University Hospital between 2012 and 2021. Tumor budding was evaluated using the International Tumor Budding Consensus Conference scoring system. Tumor budding was classified into low (0-4), intermediate (5-9), and high (≥10). For statistical analysis, tumor budding was divided into two groups based on the cut-off value of 10 (lower: 0-9 vs. higher: ≥10). The correlations between clinicopathological parameters were examined using the chi-square and Fisher's exact test. The prognostic values of the variables were analyzed using the log-rank test and Cox regression analysis. Results: Low, intermediate, and high tumor buddings were identified in 135 (61.6%), 63 (28.8), and 21 (9.6%), patients, respectively. Higher tumor budding was related to the presence of lymphatic invasion (p = 0.017), higher tumor grade (p = 0.001), higher N category (p = 0.034). In the univariable and multivariable analyses, higher tumor budding was associated with shorter disease-free survival in 97 (44.3%) patients who underwent R0 resection (p < 0.001 and p = 0.011). Tumor budding did not significantly correlate with disease-specific survival in entire patients. Conclusion: Tumor budding may serve as a prognostic factor for intrahepatic and extrahepatic cholangiocarcinomas treated with R0 resection.

Comparative proteome profiling of susceptible and resistant rice cultivars identified an arginase involved in rice defense against Xanthomonas oryzae pv. oryzae.

Xanthomonas oryzae pv. oryzae (Xoo), the causative agent of bacterial blight, is one of the major threats to rice productivity. Yet, the molecular mechanism of rice-Xoo interaction is elusive. Here, we report comparative proteome profiles of Xoo susceptible (Dongjin) and resistant (Hwayeong) cultivars of rice in response to two-time points (3 and 6 days) of Xoo infection. Low-abundance proteins were enriched using a protamine sulfate (PS) precipitation method and isolated proteins were quantified by a label-free quantitative analysis, leading to the identification of 3846 proteins. Of these, 1128 proteins were significantly changed between mock and Xoo infected plants of Dongjin and Hwayeong cultivars. Based on the abundance pattern and functions of the identified proteins, a total of 23 candidate proteins were shortlisted that potentially participate in plant defense against Xoo in the resistant cultivar. Of these candidate proteins, a mitochondrial arginase-1 showed Hwayeong specific abundance and was significantly accumulated following Xoo inoculation. Overexpression of arginase 1 (OsArg 1) in susceptible rice cultivar (Dongjin) resulted in enhanced tolerance against Xoo as compared to the wild-type. In addition, expression analysis of defense-related genes encoding PR1, glucanase I, and chitinase II by qRT-PCR showed their enhanced expression in the overexpression lines as compared to wild-type. Taken together, our results uncover the proteome changes in the rice cultivars and highlight the functions of OsARG1 in plant defense against Xoo.

Mutations in OsDET1, OsCOP10, and OsDDB1 confer embryonic lethality and alter flavonoid accumulation in Rice (Oryza sativa L.) seed.

Morphological and biochemical changes accompanying embryogenesis and seed development are crucial for plant survival and crop productivity. Here, we identified a novel yellowish-pericarp embryo lethal (yel) mutant of the japonica rice cultivar Sindongjin (Oryza sativa L.), namely, yel-sdj. Seeds of the yel-sdj mutant showed a yellowish pericarp and black embryo, and were embryonic lethal. Compared with wild-type seeds, the yel-sdj mutant seeds exhibited significantly reduced grain size, grain weight, and embryo weight, and a remarkably lower rate of embryo retention in kernels subjected to milling. However, the volume of air space between embryo and endosperm, density of embryo, and total phenolic content (TPC) and antioxidant activity of mature grains were significantly higher in the yel-sdj mutant than in the wild type. Genetic analysis and mapping revealed that the yel-sdj mutant was non-allelic to the oscop1 null mutants yel-hc, yel-cc, and yel-sk, and its phenotype was controlled by a single recessive gene, LOC_Os01g01484, an ortholog of Arabidopsis thaliana DE-ETIOLATED 1 (DET1). The yel-sdj mutant carried a 7 bp deletion in the second exon of OsDET1. Seeds of the osdet1 knockout mutant, generated via CRISPR/Cas9-based gene editing, displayed the yel mutant phenotype. Consistent with the fact that OsDET1 interacts with CONSTITUTIVE PHOTOMORPHOGENIC 10 (OsCOP10) and UV-DAMAGED DNA BINDING PROTEIN 1 (OsDDB1) to form the COP10-DET1-DDB1 (CDD), seeds of oscop10 and osddb1 knockout mutants also showed the yel phenotype. These findings will enhance our understanding of the functional roles of OsDET1 and the CDD complex in embryogenesis and flavonoid biosynthesis in rice seeds.

Speed vernalization to accelerate generation advance in winter cereal crops.

There are many challenges facing the development of high-yielding, nutritious crops for future environments. One limiting factor is generation time, which prolongs research and plant breeding timelines. Recent advances in speed breeding protocols have dramatically reduced generation time for many short-day and long-day species by optimizing light and temperature conditions during plant growth. However, winter crops with a vernalization requirement still require up to 6-10 weeks in low-temperature conditions before the transition to reproductive development. Here, we tested a suite of environmental conditions and protocols to investigate whether the vernalization process can be accelerated. We identified a vernalization method consisting of exposing seeds at the soil surface to an extended photoperiod of 22 h day:2 h night at 10°C with transfer to speed breeding conditions that dramatically reduces generation time in both winter wheat (Triticum aestivum) and winter barley (Hordeum vulgare). Implementation of the speed vernalization protocol followed by speed breeding allowed the completion of up to five generations per year for winter wheat or barley, whereas only two generations can be typically completed under standard vernalization and plant growth conditions. The speed vernalization protocol developed in this study has great potential to accelerate biological research and breeding outcomes for winter crops.

TMT-based quantitative membrane proteomics identified PRRs potentially involved in the perception of MSP1 in rice leaves.

Pathogen-associated molecular patterns (PAMPs) play a key role in triggering PAMPs triggered immunity (PTI) in plants. In the case of the rice-Magnaporthe oryzae pathosystem, fewer PAMPs and their pattern recognition receptors (PRRs) have been characterized. Recently, a M. oryzae snodprot1 homolog protein (MSP1) has been identified that functions as PAMP and triggering the PTI responses in rice. However, the molecular mechanism underlying MSP1-induced PTI is currently elusive. Therefore, we generated MSP1 overexpressed transgenic lines of rice, and a tandem mass tag (TMT)-based quantitative membrane proteomic analysis was employed to decipher the potential MSP1-induced signaling in rice using total cytosolic as well as membrane protein fractions. This approach led to the identification of 8033 proteins of which 1826 were differentially modulated in response to overexpression of MSP1 and/or exogenous jasmonic acid treatment. Of these, 20 plasma membrane-localized receptor-like kinases (RLKs) showed increased abundance in MSP1 overexpression lines. Moreover, activation of proteins related to the protein degradation and modification, calcium signaling, redox, and MAPK signaling was observed in transgenic lines expressing MSP1 in the apoplast. Taken together, our results identified potential PRR candidates involved in MSP1 recognition and suggested the overview mechanism of the MSP1-induced PTI signaling in rice leaves. SIGNIFICANCE: In plants, recognition of pathogen pathogen-derived molecules, such as PAMPs, by plant plant-derived PRRs has an essential role for in the activation of PTI against pathogen invasion. Typically, PAMPs are recognized by plasma membrane (PM) localized PRRs, however, identifying the PM-localized PRR proteins is challenging due to their low abundance. In this study, we performed an integrated membrane protein enrichment by microsomal membrane extraction (MME) method and subsequent TMT-labeling-based quantitative proteomic analysis using MSP1 overexpressed rice. Based on these results, we successfully identified various intracellular and membrane membrane-localized proteins that participated in the MSP1-induced immune response and characterized the potential PM-localized PRR candidates in rice.

Multi-omics analysis reveals the genetic basis of rice fragrance mediated by betaine aldehyde dehydrogenase 2.

INTRODUCTION: Fragrance is an important economic and quality trait in rice. The trait is controlled by the recessive gene betaine aldehyde dehydrogenase 2 (BADH2) via the production of 2-acetyl-1-pyrroline (2AP). OBJECTIVES: Variation in BADH2 was evaluated at the population, genetic, transcriptional, and metabolic levels to obtain insights into fragrance regulation in rice. METHODS: Whole-genome resequencing of the Korean World Rice Collection of 475 rice accessions, including 421 breeding lines and 54 wild accessions, was performed. Transcriptome analyses of a subset of 279 accessions, proteome analyses of 64 accessions, and volatile profiling of 421 breeding lines were also performed. RESULTS: We identified over 3.1 million high-quality single nucleotide polymorphisms (SNPs) in Korean rice collection. Most SNPs were present in intergenic regions (79%), and 190,148 SNPs (6%) were located in the coding sequence, of which 53% were nonsynonymous. In total, 38 haplotypes were identified in the BADH2 coding region, including four novel haplotypes (one in cultivated and three in wild accessions). Tajima's D values suggested that BADH2 was under balancing selection in japonica rice. Furthermore, we identified 316 expression quantitative trait loci (eQTL), including 185 cis-eQTLs and 131 trans-eQTLs, involved in BADH2 regulation. A protein quantitative trait loci (pQTL) analysis revealed the presence of trans-pQTLs; 13 pQTLs were mapped 1 Mbp from the BADH2 region. Based on variable importance in projection (VIP) scores, 15 volatile compounds, including 2AP, discriminated haplotypes and were potential biomarkers for rice fragrance. CONCLUSION: We generated a catalog of haplotypes based on a resequencing analysis of a large number of rice accessions. eQTLs and pQTLs associated with BADH2 gene expression and protein accumulation are likely involved in the regulation of 2AP variation in fragrant rice. These data improve our understanding of fragrance and provide valuable information for rice breeding.

Shotgun proteomic analysis for detecting differentially expressed proteins in the reduced culm number rice.

To survey protein expression patterns in the reduced culm number (RCN) rice, a comparative shotgun proteomic analysis was conducted. For large-scale protein identification, multidimensional protein identification technology (MudPIT) coupled with pre-fractionation of plant shoot proteins led to the identification of 3004 non-redundant rice proteins. By statistically comparing relative amounts of 1353 reproducibly identified proteins between the RCN rice and the wild-type rice, 44 differentially expressed proteins were detected, where 42 proteins were increased and 2 proteins were decreased in the RCN rice. These proteins appear to have roles in glycolysis, trichloroacetic acid cycle, secondary metabolism, nutrient recycling, and nucleotide metabolism and repair. Consequently, we hypothesized that the RCN rice might fail to maintain sugar nutrient homeostasis. This was confirmed with the observation that the sucrose concentration was increased significantly in the RCN rice compared with the wild-type rice. Also, the RCN rice showed a hypersensitive response to exogenous sucrose treatment.

Genome-Wide Identification and Genetic Variations of the Starch Synthase Gene Family in Rice.

Starch is a major ingredient in rice, and the amylose content of starch significantly impacts rice quality. OsSS (starch synthase) is a gene family related to the synthesis of amylose and amylopectin, and 10 members have been reported. In the present study, a synteny analysis of a novel family member belonging to the OsSSIV subfamily that contained a starch synthase catalytic domain showed that three segmental duplications and multiple duplications were identified in rice and other species. Expression data showed that the OsSS gene family is involved in diverse expression patterns. The prediction of miRNA targets suggested that OsSS are possibly widely regulated by miRNA functions, with miR156s targeted to OsSSII-3, especially. Haplotype analysis exhibited the relationship between amylose content and diverse genotypes. These results give new insight and a theoretical basis for the improved amylose content and eating quality of rice.

Genome-Wide Association Study Reveals the Genetic Basis of Chilling Tolerance in Rice at the Reproductive Stage.

A genome-wide association study (GWAS) was used to investigate the genetic basis of chilling tolerance in a collection of 117 rice accessions, including 26 Korean landraces and 29 weedy rices, at the reproductive stage. To assess chilling tolerance at the early young microspore stage, plants were treated at 12 °C for 5 days, and tolerance was evaluated using seed set fertility. GWAS, together with principal component analysis and kinship matrix analysis, revealed five quantitative trait loci (QTLs) associated with chilling tolerance on chromosomes 3, 6, and 7. The percentage of phenotypic variation explained by the QTLs was 11-19%. The genomic region underlying the QTL on chromosome 3 overlapped with a previously reported QTL associated with spikelet fertility. Subsequent bioinformatic and haplotype analyses suggested three candidate chilling-tolerance genes within the QTL linkage disequilibrium block: Os03g0305700, encoding a protein similar to peptide chain release factor 2; Os06g0495700, encoding a beta tubulin, autoregulation binding-site-domain-containing protein; and Os07g0137800, encoding a protein kinase, core-domain-containing protein. Further analysis of the detected QTLs and the candidate chilling-tolerance genes will facilitate strategies for developing chilling-tolerant rice cultivars in breeding programs.

OsGATA16, a GATA Transcription Factor, Confers Cold Tolerance by Repressing OsWRKY45-1 at the Seedling Stage in Rice.

BACKGROUND: Cold stress is the main abiotic stress in rice, which seriously affects the growth and yield of rice. Identification of cold tolerance genes is of great significance for rice to solve these problems. GATA-family transcription factors involve diverse biological functions, however, their role in cold tolerance in rice remains unclear. RESULTS: In this study, a GATA-type zinc finger transcription factor OsGATA16, which can improve cold tolerance, was isolated and characterized from rice. OsGATA16 belongs to OsGATA subfamily-II and contains 11 putative phosphorylation sites, a nuclear localization signal (NLS), and other several conserved domains. OsGATA16 was expressed in all plant tissues, with the strongest in panicles. It was induced by cold and ABA treatments, but was repressed by drought, cytokinin and JA, and acted as a transcriptional suppressor in the nucleus. Overexpression of OsGATA16 improves cold tolerance of rice at seedling stage. Under cold stress treatments, the transcription of four cold-related genes OsWRKY45-1, OsSRFP1, OsCYL4, and OsMYB30 was repressed in OsGATA16-overexpressing (OE) rice compared with wild-type (WT). Interestingly, OsGATA16 bound to the promoter of OsWRKY45-1 and repressed its expression. In addition, haplotype analysis showed that OsGATA16 polarized between the two major rice subspecies japonica and indica, and had a non-synonymous SNP8 (336G) associated with cold tolerance. CONCLUSION: OsGATA16 is a GATA transcription factor, which improves cold tolerance at seedling stage in rice. It acts as a positive regulator of cold tolerance by repressing some cold-related genes such as OsWRKY45-1, OsSRFP1, OsCYL4 and OsMYB30. Additionally, OsGATA16 has a non-synonymous SNP8 (336G) associated with cold tolerance on CDS region. This study provides a theoretical basis for elucidating the mechanism of cold tolerance in rice and new germplasm resources for rice breeding.