Lijun Luo

Natural variation in GS5 plays an important role in regulating grain size and yield in rice

Increasing crop yield is one of the most important goals of plant science research. Grain size is a major determinant of grain yield in cereals and is a target trait for both domestication and artificial breeding. We showed that the quantitative trait locus (QTL) GS5 in rice controls grain size by regulating grain width, filling and weight. GS5 encodes a putative serine carboxypeptidase and functions as a positive regulator of grain size, such that higher expression of GS5 is correlated with larger grain size. Sequencing of the promoter region in 51 rice accessions from a wide geographic range identified three haplotypes that seem to be associated with grain width. The results suggest that natural variation in GS5 contributes to grain size diversity in rice and may be useful in improving yield in rice and, potentially, other crops.

Genetic analysis for drought resistance of rice at reproductive stage in field with different types of soil

Drought resistance of rice is a complex trait and is mainly determined by mechanisms of drought avoidance and drought tolerance. The present study was conducted to characterize the genetic basis of drought resistance at reproductive stage in field by analyzing the QTLs for drought response index (DRI, normalized by potential yield and flowering time), relative yield, relative spikelet fertility, and four traits of plant water status and their relationships with root traits using a recombinant inbred population derived from a cross between an indica rice and upland rice. A total of 39 QTLs for these traits were detected with individual QTL explained 5.1–32.1% of phenotypic variation. Only two QTLs for plant water status were commonly detected in two environments, suggesting different mechanisms might exist in two types of soil conditions. DRI has no correlation with potential yield and flowering time under control, suggesting that it can be used as a good drought resistance index in field conditions. The co-location of QTLs for canopy temperature and delaying in flowering time suggested a usefulness of these two traits as indexes in drought resistance screening. Correlation and QTL congruence between root traits and putative drought tolerance traits revealed that drought avoidance (via thick and deep root traits) was the main genetic basis of drought resistance in sandy soil condition, while drought tolerance may play more role in the genetic basis of drought resistance in paddy soil condition. Therefore, both drought mechanisms and soil textures must be considered in the improvement of drought resistance at reproductive stage in rice.

Chalk5 encodes a vacuolar H+-translocating pyrophosphatase influencing grain chalkiness in rice

Grain chalkiness is a highly undesirable quality trait in the marketing and consumption of rice grain. However, the molecular basis of this trait is poorly understood. Here we show that a major quantitative trait locus (QTL), Chalk5, influences grain chalkiness, which also affects head rice yield and many other quality traits. Chalk5 encodes a vacuolar H+-translocating pyrophosphatase (V-PPase) with inorganic pyrophosphate (PPi) hydrolysis and H+-translocation activity. Elevated expression of Chalk5 increases the chalkiness of the endosperm, putatively by disturbing the pH homeostasis of the endomembrane trafficking system in developing seeds, which affects the biogenesis of protein bodies and is coupled with a great increase in small vesicle-like structures, thus forming air spaces among endosperm storage substances and resulting in chalky grain. Our results indicate that two consensus nucleotide polymorphisms in the Chalk5 promoter in rice varieties might partly account for the differences in Chalk5 mRNA levels that contribute to natural variation in grain chalkiness.

Quantitative trait loci for panicle size, heading date and plant height co-segregating in trait-performance derived near-isogenic lines of rice ( Oryza sativa )

Near-isogenic lines (NILs) are ideal materials for precise estimation of quantitative trait loci (QTL) effects and map-based gene isolation. With the completion of the rice genome sequence, QTL isolation based on NILs is becoming a routine. In this study, a trait-performance derived NIL strategy was adopted to develop NILs. Two plants were identified within one inbred line of recombinant inbred lines (RILs, F7 generation), exhibiting a significant difference in panicle size. By marker screening of the whole genome the genetic background of the two plants was estimated to be 98.7% identical. These two plants were selected as parents to produce a near-isogenic F2 (NIL-F2) population, consisting of 125 individuals, in which spikelets per panicle (SPP), grains per panicle (GPP), heading date (HD) and plant height (PH) were recorded. These four traits expressed discontinuous or bimodal distribution in the NIL-F2 population and followed the expected segregation ratios for a single Mendelian factor by progeny tests. A partial dominant QTL for the four traits was mapped to the same interval flanked by RM310 and RM126 on chromosome 8. The QTL region explained 83.0, 80.2, 94.9 and 93.8% of trait variation of SPP, GPP, HD and PH in the progenies, respectively. Progeny tests also confirmed co-segregation of QTL for the four traits, tall plants consistently flowering late and carrying large panicles. Different NILs development strategies are discussed.

Genetic, proteomic and metabolic analysis of the regulation of energy storage in rice seedlings in response to drought

We used proteomic analysis to determine the response of rice plant seedlings to drought‐induced stress. The expression of 71 protein spots was significantly altered, and 60 spots were successfully identified. The greatest down‐regulated protein functional category was translation. Up‐regulated proteins were mainly related to protein folding and assembly. Additionally, many proteins involved in metabolism (e.g. carbohydrate metabolism) also showed differences in expression. cDNA microarray and GC‐MS analysis showed 4756 differentially expressed mRNAs and 37 differentially expressed metabolites. Once these data were integrated with the proteomic analysis, we were able to elucidate the metabolic pathways affected by drought‐induced stress. These results suggest that increased energy consumption from storage substances occurred during drought. In addition, increased expression of the enzymes involved in anabolic pathways corresponded with an increase in the content of six amino acids. We speculated that energy conversion from carbohydrates and/or fatty acids to amino acids was increased. Analysis of basic metabolism networks allowed us to understand how rice plants adjust to drought conditions.

OsGRAS23 , a rice GRAS transcription factor gene, is involved in drought stress response through regulating expression of stress-responsive genes

BackgroundDrought is a major abiotic stress factors that reduces agricultural productivity. GRAS transcription factors are plant-specific proteins that play diverse roles in plant development. However, the functions of a number of GRAS genes identified in rice are unknown, especially the GRAS genes related to rice drought resistance have not been characterized.ResultsIn this study, a novel GRAS transcription factor gene named OsGRAS23, which is located in a drought-resistant QTL interval on chromosome 4 of rice, was isolated. The expression of OsGRAS23 was induced by drought, NaCl, and jasmonic acid treatments. The OsGRAS23-GFP fused protein was localized in the nucleus of tobacco epidermal cells. A trans-activation assay in yeast cells demonstrated that the OsGRAS23 protein possessed a strong transcriptional activation activity. OsGRAS23-overexpressing rice plants showed improved drought resistance and oxidative stress tolerance as well as less H2O2 accumulation compared with the wild-type plants. Furthermore, microarray analysis showed that several anti-oxidation related genes were up-regulated in the OsGRAS23-overexpressing rice plants. The yeast one hybrid test indicated that OsGRAS23 could bind to the promoters of its potential target genes.ConclusionsOur results demonstrate that OsGRAS23 encodes a stress-responsive GRAS transcription factor and positively modulates rice drought tolerance via the induction of a number of stress-responsive genes.

Correlation analysis and QTL identification for canopy temperature, leaf water potential and spikelet fertility in rice under contrasting moisture regimes

Canopy temperature (CT), leaf water potential (LWP) and spikelet fertility (SF) of a set of RILs (F9) from the cross between Zhenshan97B and IRAT109 were investigated under two soil moisture regimes in a drought screen facility. In water stress condition, CT was negatively correlated with SF (r = −0.2867) and LWP (r = −0.2740), and LWP was positively associated with SF (r = 0.1696). These results indicated that the plant with high drought tolerant ability could maintain higher LWP and lower CT, leading to higher SF. A total of 44 main effect QTLs were associated with CT, LWP and SF. The accumulated contributions of QTLs for CT, LWP and SF were 87.85%, 15.06%, 79.46% under well water condition and 72.61%, 87.68%, 33.29% under stress condition, respectively. Totally 45 pairs of digenic interactions were detected. The accumulated contribution of digenic epistasis on CT, LWP and SF were 55.69%, 47.15%, 48.15% under well water condition and 53.44%, 57.94%, 54.62% under water stress condition. Compared with other drought tolerance QTL mapping researches in rice, 19 main effect QTLs were found to be located at the same or closely conjoint regions.

Transgenerational Variations in DNA Methylation Induced by Drought Stress in Two Rice Varieties with Distinguished Difference to Drought Resistance

Adverse environmental conditions have large impacts on plant growth and crop production. One of the crucial mechanisms that plants use in variable and stressful natural environments is gene expression modulation through epigenetic modification. In this study, two rice varieties with different drought resistance levels were cultivated under drought stress from tilling stage to seed filling stage for six successive generations. The variations in DNA methylation of the original generation (G0) and the sixth generation (G6) of these two varieties in normal condition (CK) and under drought stress (DT) at seedling stage were assessed by using Methylation Sensitive Amplification Polymorphism (MSAP) method. The results revealed that drought stress had a cumulative effect on the DNA methylation pattern of both varieties, but these two varieties had different responses to drought stress in DNA methylation. The DNA methylation levels of II-32B (sensitive) and Huhan-3 (resistant) were around 39% and 32%, respectively. Genome-wide DNA methylation variations among generations or treatments accounted for around 13.1% of total MSAP loci in II-32B, but was only approximately 1.3% in Huhan-3. In II-32B, 27.6% of total differentially methylated loci (DML) were directly induced by drought stress and 3.2% of total DML stably transmitted their changed DNA methylation status to the next generation. In Huhan-3, the numbers were 48.8% and 29.8%, respectively. Therefore, entrainment had greater effect on Huhan-3 than on II-32B. Sequence analysis revealed that the DML were widely distributed on all 12 rice chromosomes and that it mainly occurred on the gene’s promoter and exon region. Some genes with DML respond to environmental stresses. The inheritance of epigenetic variations induced by drought stress may provide a new way to develop drought resistant rice varieties.

Identification of Rice Transcription Factors Associated with Drought Tolerance Using the Ecotilling Method

The drought tolerance (DT) of plants is a complex quantitative trait. Under natural and artificial selection, drought tolerance represents the crop survival ability and production capacity under drought conditions (Luo, 2010). To understand the regulation mechanism of varied drought tolerance among rice genotypes, 95 diverse rice landraces or varieties were evaluated within a field screen facility based on the ‘line–source soil moisture gradient’, and their resistance varied from extremely resistant to sensitive. The method of Ecotype Targeting Induced Local Lesions in Genomes (Ecotilling) was used to analyze the diversity in the promoters of 24 transcription factor families. The bands separated by electrophoresis using Ecotilling were converted into molecular markers. STRUCTURE analysis revealed a value of K = 2, namely, the population with two subgroups (i.e., indica and japonica), which coincided very well with the UPGMA clusters (NTSYS-pc software) using distance-based analysis and InDel markers. Then the association analysis between the promoter diversity of these transcription factors and the DT index/level of each variety was performed. The results showed that three genes were associated with the DT index and that five genes were associated with the DT level. The sequences of these associated genes are complex and variable, especially at approximately 1000 bp upstream of the transcription initiation sites. The study illuminated that association analysis aimed at Ecotilling diversity of natural groups could facilitate the isolation of rice genes related to complex quantitative traits.

The QTL controlling amino acid content in grains of rice ( Oryza sativa ) are co-localized with the regions involved in the amino acid metabolism pathway

The improvement of grain quality, such as protein content (PC) and amino acid composition, has been a major concern of rice breeders. We constructed a population of 190 recombinant inbred lines (RILs) from a cross between Zhenshan 97 and Nanyangzhan to map the quantitative trait locus or loci (QTL) for amino acid content (AAC) as characterized by each of the AACs, total essential AAC, and all AAC. Using the data collected from milled rice in 2002 and 2004, we identified 18 chromosomal regions for 19 components of AAC. For 13 of all the loci, the Zhenshan 97 allele increased the trait values. Most QTL were co-localized, forming ten QTL clusters in 2002 and six in 2004. The QTL clusters varied in both effects and locations, and the mean values of variation explained by individual QTL in the clusters ranged from 4.3% to 28.82%. A relatively strong QTL cluster, consisting of up to 19 individual QTL, was found at the bottom of chromosome 1. The major QTL clusters identified for two different years were coincident. A wide coincidence was found between the QTL we detected and the loci involved in amino acid metabolism pathways, including N assimilation and transfer, and amino acid or protein biosynthesis. The results will be useful for candidate gene identification and marker-assisted favorable allele transfer in rice breeding programs.