RI Lines Research Articles

Molecular and cytogenetic characterization of Osteospermum fruticosum lines harboring wild type pRi rol genes.

Transgenic lines engineered through wild type Rhizobium rhizogenes display an altered phenotype known as the Ri phenotype. This phenotype includes a more compact plant habit, which has proved useful to obtain more compact varieties that require less chemical growth regulation. Here, we develop a method for the molecular and cytogenetic characterization of Cape daisy (Osteospermum fruticosum Norl.) Ri lines in order to predict segregation of pRi T-DNA genes. Analysis of copy number variation (CNV) by means of digital PCR indicated large variation in the copy number of the inserted root oncogenic loci (rol) genes, ranging from 1 to more than 15 copies. In addition, up to 9 copies of the auxin biosynthesis genes (aux) were present in a single Ri line. Visualization of pRiA4 and pRi1724 rol and aux insertion in 4 Ri lines was performed through Fluorescence In Situ Hybridization. The number of rol integrated loci varied from 1 to 3 loci. In contrast, the different TR-gene copies were confined to a single locus which consistently co-localized with a TL locus, this was demonstrated for the first time. Based on CNV and FISH a single Ri line, harboring 7 pRi1724 rol gene copies dispersed over 3 integration loci, was selected for breeding. Copy number segregation in R1 progeny of 2, 3, 4 and 5 pRi1724 copies was confirmed, indicating that the evaluation of the breeding value of first generation Ri lines is possible through CNV and FISH.

Molecular and biochemical components associated with chilling tolerance in tomato: comparison of different developmental stages

The cultivated tomato, Solanum lycopersicum, is highly sensitive to cold stress (CS), resulting in significant losses during cultivation and postharvest fruit storage. Previously, we demonstrated the presence of substantial genetic variation in fruit chilling tolerance in a tomato recombinant inbred line (RIL) population derived from a cross between a chilling-sensitive tomato line and a chilling-tolerant accession of the wild species S. pimpinellifolium. Here, we investigated molecular and biochemical components associated with chilling tolerance in fruit and leaves, using contrasting groups of “chilling tolerant” and “chilling sensitive” RI lines. Transcriptomic analyses were conducted on fruit exposed to CS, and gene expressions and biochemical components were measured in fruit and leaves. The analyses revealed core responding genes specific to either the cold-tolerant or cold-sensitive RI lines, which were differentially regulated in similar fashion in both leaves and fruit within each group. These genes may be used as markers to determine tomato germplasm cold tolerance or sensitivity. This study demonstrated that tomato response to CS in different developmental stages, including seedling and postharvest fruit, might be mediated by common biological/genetic factors. Therefore, genetic selection for cold tolerance during early stages of plant development may lead to lines with greater postharvest fruit chilling tolerance.

Superior osmotic stress tolerance in oilseed rape transformed with wild-type Rhizobium rhizogenes

Key messageNatural transformation with R. rhizogenes enhances osmotic stress tolerance in oilseed rape through increasing osmoregulation capacity, enhancing maintenance of hydraulic integrity and total antioxidant capacity.Transformation of plants using wild strains of agrobacteria is termed natural transformation and is not covered by GMO legislation in, e.g., European Union and Japan. In this study, offspring lines of Rhizobium rhizogenes naturally transformed oilseed rape (Brassica napus), i.e., A11 and B3 (termed root-inducing (Ri) lines), were investigated for osmotic stress resilience. Under polyethylene glycol 6000 (PEG) 10% (w/v)-induced osmotic stress, the Ri lines, particularly A11, had less severe leaf wilting, higher stomatal conductance (8.2 times more than WT), and a stable leaf transpiration rate (about 2.9 mmol m−2 s−1). Although the leaf relative water content and leaf water potential responded similarly to PEG treatment between the Ri lines and WT, a significant reduction of the turgid weight to dry weight ratio in A11 and B3 indicated a greater capacity of osmoregulation in the Ri lines. Moreover, the upregulation of plasma membrane intrinsic proteins genes (PIPs) in roots and downregulation of these genes in leaves of the Ri lines implied a better maintenance of hydraulic integrity in relation to the WT. Furthermore, the Ri lines had greater total antioxidant capacity (TAC) than the WT under PEG stress. Collectively, the enhanced tolerance of the Ri lines to PEG-induced osmotic stress could be attributed to the greater osmoregulation capacity, better maintenance of hydraulic integrity, and greater TAC than the WT. In addition, Ri-genes (particularly rolA and rolD) play roles in response to osmotic stress in Ri oilseed rape. This study reveals the potential of R. rhizogenes transformation for application in plant drought resilience.

Optimization of Rhizobium rhizogenes-mediated transformation, regeneration and characterization of Malus domesticaBorkh. Ri lines

This study explored the establishment and optimization of Ri (root inducing) technology for apple breeding, using the bacterium Rhizobium rhizogenes to obtain Ri lines with compact shoots and stronger root systems. The transformation and shoot regeneration for Malus domestica cultivars was studied in detail. Various R. rhizogenes strains, scion and rootstock genotypes, explant types, wounding methods and explant orientations were tested for hairy root induction. Most of the 16 tested strains, especially those with plasmid type III, induced hairy roots in the rootstock genotype ‘M26’. Although apple genotypes differed in response, in most of them roots were successfully induced using strains ATCC 15834, LMG 63 and LMG 150, with leaf blades outperforming petioles as explants. Wounding by scratching or sonication further improved transformation efficiency, as did placing leaf blades with their abaxial side upward on root induction medium. The majority (94%) of roots formed in one transformation experiment were tested PCR-positive for at least one T-DNA gene. Shoot regeneration experiments investigated salt concentrations, gelling agents, cytokinin types, concentrations, and a resting period on hormone-free medium. Shoot regeneration was highly genotype-dependent varying between 0 and 83%, whereas only minor, non-significant effects were observed for the treatments tested. Copy numbers of T-DNA genes were estimated using digital PCR for the first time in apple Ri lines. In the greenhouse, two Ri lines showed compact shoots and shorter leaves, but no enhanced root system. The improved protocol provides a valuable tool for breeders and scientists to obtain and further use Ri lines.

Agrobacterium rhizogenes: paving the road to research and breeding for woody plants.

Woody plants play a vital role in global ecosystems and serve as valuable resources for various industries and human needs. While many woody plant genomes have been fully sequenced, gene function research and biotechnological breeding advances have lagged behind. As a result, only a limited number of genes have been elucidated, making it difficult to use newer tools such as CRISPR-Cas9 for biotechnological breeding purposes. The use of Agrobacterium rhizogenes as a transformative tool in plant biotechnology has received considerable attention in recent years, particularly in the research field on woody plants. Over the past three decades, numerous woody plants have been effectively transformed using A. rhizogenes-mediated techniques. Some of these transformed plants have successfully regenerated. Recent research on A. rhizogenes-mediated transformation of woody plants has demonstrated its potential for various applications, including gene function analysis, gene expression profiling, gene interaction studies, and gene regulation analysis. The introduction of the Ri plasmid has resulted in the emergence of several Ri phenotypes, such as compact plant types, which can be exploited for Ri breeding purposes. This review paper presents recent advances in A. rhizogenes-mediated basic research and Ri breeding in woody plants. This study highlights various aspects of A. rhizogenes-mediated transformation, its multiple applications in gene function analysis, and the potential of Ri lines as valuable breeding materials.

Improved compact growth habit of Viola × wittrockiana through Rhizobium rhizogenes transformation

Compactness is an important breeding goal in the improvement of garden pansy varieties (Viola × wittrockiana Gams.). The desired compact stature is primarily obtained through the use of chemical growth retardants. Recently, an alternative strategy for creating compact ornamentals was proposed based on transformation by wild type rhizogenic agrobacteria and the regeneration of transformed hairy roots. Here, we report on the successful transformation and creation of compact garden pansy lines. The most efficient hairy root induction was obtained by Rhizobium rhizogenes strain ATCC15834. Shoot regeneration from hairy roots was obtained using 3 mg L−1 2iP and 0.3 mg L−1 2,4-D. The presence of R. rhizogenes T-DNA genes was confirmed in the regenerated lines. These lines displayed a varying degrees of dwarfed growth. Most notably the shoot length was reduced up to 49%. Other phenotypic traits that were induced include reduced peduncle length and smaller, wrinkled flowers. Several of these Ri lines with promising compactness have been selected for further breeding towards compact garden pansy varieties that are less dependent on chemical growth regulation.Graphical abstract

Reevaluation of the effect of dietary restriction on different recombinant inbred lines of male and female mice.

Dietary restriction (DR) was reported to either have no effect or reduce the lifespan of the majority of the 41‐recombinant inbred (RI) lines studied by Liao et al. (Aging Cell, 2010, 9, 92). In an appropriately power longevity study (n > 30 mice/group), we measured the lifespan of the four RI lines (115‐RI, 97‐RI, 98‐RI, and 107‐RI) that were reported to have the greatest decrease in lifespan when fed 40% DR. DR increased the median lifespan of female RI‐115, 97‐RI, and 107‐RI mice and male 115‐RI mice. DR had little effect (<4%) on the median lifespan of female and male 98‐RI mice and male 97‐RI mice and reduced the lifespan of male 107‐RI mice over 20%. While our study was unable to replicate the effect of DR on the lifespan of the RI mice (except male 107‐RI mice) reported by Liao et al. (Aging Cell, 2010, 9, 92), we found that the genotype of a mouse had a major impact on the effect of DR on lifespan, with the effect of DR ranging from a 50% increase to a 22% decrease in median lifespan. No correlation was observed between the changes in either body composition or glucose tolerance induced by DR and the changes observed in lifespan of the four RI lines of male and female mice. These four RI lines of mice give the research community a unique resource where investigators for the first time can study the anti‐aging mechanism of DR by comparing mice in which DR increases lifespan to mice where DR has either no effect or reduces lifespan.

Compact shoot architecture of Osteospermum fruticosum transformed with Rhizobium rhizogenes.

Improved compact shoot architecture of Osteospermum fruticosum Ri lines obtained through Rhizobium rhizogenes transformation reduces the need for chemical growth retardants. Compactness is for many ornamental crops an important commercial trait that is usually obtained through the application of growth retardants. Here, we have adopted a genetic strategy to introduce compactness in the perennial shrub Cape daisy (Osteospermum fruticosum Norl.). To this end, O. fruticosum was transformed using six different wild type Rhizobium rhizogenes strains. The most effective R. rhizogenes strains Arqua1 and ATCC15834 were used to create hairy root cultures from six Cape daisy genotypes. These root cultures were regenerated to produce transgenic Ri lines, which were analyzed for compactness. Ri lines displayed the characteristic Ri phenotype, i.e., reduced plant height, increased branching, shortened internodes, shortened peduncles, and smaller flowers. Evaluation of the Ri lines under commercial production conditions showed that similar compactness was obtained as the original Cape daisy genotypes treated with growth retardant. The results suggest that the use of chemical growth retardants may be omitted or reduced in commercial production systems of Cape daisy through implementation of Ri lines in future breeding programs.

RNAi silenced ζ-carotene desaturase developed variegated tomato transformants with increased phytoene content

ζ-carotene desaturase (ZDS) is one of the enzymes that catalyzes the conversion of ζ-carotene to lycopene. However, the role of ZDS has been not studied extensively in tomato. In this study, to examine the role of ZDS in tomato, the expression of ZDS was downregulated by RNA interference. The relative mRNA levels of ZDS in RNAi lines were reduced by more than 75% compared to wild-type plants. The genetically modified plants exhibited altered morphological characteristics such as photobleached leaves, increased leaf length, longer petioles, larger leaf surface areas and whitish-green tomato fruits. Chromatographic determination showed a significant increase in phytoene in both leaves and fruits of the RI lines, whereas other types of carotenoids were significantly decreased compared to controls. qPCR analysis revealed significant changes in the expression of PSY, PDS and LCYB. The endogenous hormones and flavonoid levels were altered in leaves and fruits of ZDS-silenced plants. Electron microscopic studies revealed hampered chloroplast differentiation and disrupted thylakoid structure in the ZDS-suppressed lines. GC–MS analysis showed that the level of several organic acids, amino acids and fatty acids were decreased in the RI lines, whereas the sugars and sugar derivatives were slightly increased compared to controls. Thus, these results imply a potential for ZDS in carotenoid enhancement through genetic engineering of the carotenoid pathway.

Segregation of rol Genes in Two Generations of Sinningia speciosa Engineered Through Wild Type Rhizobium rhizogenes.

Rhizobium rhizogenes infects and transforms a wide range of plant species. It thereby introduces new genes located on transfer-DNA of the root inducing plasmid (pRi) into the plant genome and one of its abilities is to alter the host root system. Explants from pRi transformed roots from Sinningia speciosa were regenerated to create naturally transgenic Ri lines. The presence of rol and aux genes in the Ri lines was linked with altered growth characteristics: shorter peduncles, wrinkled leaves, delayed flowering and enhanced root growth. The potential of Ri lines for breeding was evaluated through consecutive backcrossing with the original host genotype. The progeny of reciprocal crosses showed non-Mendelian inheritance suggesting partial transmission of the of the aux and rol genes. The typical Ri phenotype observed in the primary Ri line was partially inherited. These results revealed that the Ri phenotype is a complex trait influenced by the genetic background of the Ri line.

Rhizogenic agrobacteria as an innovative tool for plant breeding: current achievements and limitations.

Compact plant growth is an economically important trait for many crops. In practice, compactness is frequently obtained by applying chemical plant growth regulators. In view of sustainable and environmental-friendly plant production, the search for viable alternatives is a priority for breeders. Co-cultivation and natural transformation using rhizogenic agrobacteria result in morphological alterations which together compose the Ri phenotype. This phenotype is known to exhibit a more compact plant habit, besides other features. In this review, we highlight the use of rhizogenic agrobacteria and the Ri phenotype with regard to sustainable plant production and plant breeding. An overview of described Ri lines and current breeding applications is presented. The potential of Ri lines as pre-breeding material is discussed from both a practical and legal point of view.

Thermal shift research on the R1 and R2 lines of Cr3+-doped ZnGa2O4 spinel with a full expression

This paper gives a theoretical research on the thermal shifts of Ri (i = 1, 2) lines (corresponding to the transitions from two components of the first excited state 2E to the ground state 4A2) in ZnGa2O4: Cr3+ spinel with a full expression including both the static effect due to lattice thermal expansion and the dynamic effect owing to electron-phonon interaction. The outcomes show that the static and dynamic effects result in the thermal blue- and red-shifts, respectively. The relative importances of static effect for R1 and R2 lines are, respectively, about 26% and 27% and so the observed thermal shifts of Ri lines are thermal red-shifts. The true electron-phonon coupling parameters α’ for both the Ri lines are evaluated. They are found to be about 1.36 times the corresponding apparent electron-phonon coupling parameters α acquired by considering only the dynamic effect. These results are discussed.

Rice nitrate transporter OsNPF7.2 positively regulates tiller number and grain yield

BackgroundRice tiller number is one of the most important factors that determine grain yield, while nitrogen is essential for the crop growth and development, especially for tiller formation. Genes involved in nitrogen use efficiency processes have been identified in the previous studies, however, only a small number of these genes have been found to improve grain yield by promoting tillering.ResultsWe constructed over-expression (OX) lines and RNA-interference (Ri) lines, and selected a mutant of OsNPF7.2, a low-affinity nitrate transporter. Our analyses showed that rice tiller number and grain yield were significantly increased in OX lines, whereas Ri lines and mutant osnpf7.2 had fewer tiller number and lower grain yield. Under different nitrate concentrations, tiller buds grew faster in OX lines than in WT, but they grew slower in Ri lines and mutant osnpf7.2. These results indicated that altered expression of OsNPF7.2 plays a significant role in the control of tiller bud growth and regulation of tillering. Elevated expression of OsNPF7.2 also improved root length, root number, fresh weight, and dry weight. However, reduced expression of OsNPF7.2 had the opposite result on these characters. OsNPF7.2 OX lines showed more significantly enhanced influx of nitrate and had a higher nitrate concentration than WT. The levels of gene transcripts related to cytokinin pathway and cell cycle in tiller bud, and cytokinins concentration in tiller basal portion were higher in OX lines than that in WT, suggesting that altered expression of OsNPF7.2 controlled tiller bud growth and root development by regulating cytokinins content and cell cycle in plant cells. Altered expression of OsNPF7.2 also was responsible for the change in expression of the genes involved in strigolactone pathway, such as D27, D17, D10, Os900, Os1400, D14, D3, and OsFC1.ConclusionOur results suggested that OsNPF7.2 is a positive regulator of nitrate influx and concentration, and that it also regulates cell division in tiller bud and alters expression of genes involved in cytokinin and strigolactone pathways, resulting in the control over rice tiller number. Since elevated expression of OsNPF7.2 is capable of improving rice grain yield, this gene might be applied to high-yield rice breeding.

A novel functional gene associated with cold tolerance at the seedling stage in rice.

SummaryIdentification and cloning of cold‐tolerant genes that can stably express under different cold environments are crucial for molecular rice breeding for cold tolerance. In the previous study, we identified a cold‐tolerant QTL at the seedling stage, qCTS‐9 which could be detected under different cold environments using a recombinant inbred line (RIL) population derived from a cold‐tolerant variety Lijiangxintuanheigu (LTH) and a cold‐sensitive variety Shanhuangzhan 2 (SHZ‐2). In this study, eight candidate genes within the qCTS‐9 interval were identified through integrated analysis of QTL mapping with genomewide differential expression profiling of LTH. The qRT‐PCR assay showed that only Os09g0410300 exhibited different expression patterns between LTH and SHZ‐2 during cold stress, and significantly positive correlation was found between cold induction of Os09g0410300 and seedling cold tolerance in the RI lines. Five SNPs and one InDel in the promoters of Os09g0410300 were detected between LTH and SHZ‐2, and the InDel marker ID410300 designed based on the insertion–deletion polymorphism in the promoter was significantly associated with seedling cold tolerance in RIL population. Further, Os09g0410300 over‐expression plants exhibited enhanced cold tolerance at the seedling stage compared with the wild‐type plants. Thus, our results suggest that Os09g0410300 is the functional gene underlying qCTS‐9. To our knowledge, it is a novel gene contributed to enhance cold tolerance at the seedling stage in rice. Identification of the functional gene underlying qCTS‐9 and development of the gene‐specific marker will facilitate molecular breeding for cold tolerance at the seedling stage in rice through transgenic approach and marker‐assisted selection (MAS).

A new transgenic rice line exhibiting enhanced ferric iron reduction and phytosiderophore production confers tolerance to low iron availability in calcareous soil.

Iron (Fe) deficiency is a critical agricultural problem, especially in calcareous soil, which is distributed worldwide. Rice plants take up Fe(II) from soil through a OsIRT1 transporter (Strategy I-related system) and also take up Fe(III) via a phytosiderophore-based system (Strategy II system). However, rice plants are susceptible to low-Fe conditions because they have low Fe(III) reduction activity and low-level phytosiderophore secretion. Previously, we produced transgenic rice plants expressing a mutationally reconstructed yeast ferric chelate reductase, refre1/372, under the control of the OsIRT1 promoter. This transgenic rice line exhibited higher Fe(III) chelate reductase activity and tolerance to Fe deficiency. In addition, we produced transgenic rice overexpressing the Fe deficiency-inducible transcription factor, OsIRO2, which regulates the expression of various genes involved in the strategy II Fe(III) uptake system, including OsNAS1, OsNAAT1, OsDMAS1, OsYSL15, and TOM1. This transgenic rice exhibited improved phytosiderophore secretion ability and tolerance to Fe deficiency. In the present research, transgenic rice plants that possess both the OsIRT1 promoter-refre1/372 and the 35S promoter-OsIRO2 (RI lines) were produced to enhance both Strategy I Fe(II) reductase ability and Strategy II phytosiderophore productivity. RI lines exhibited enhanced tolerance to Fe-deficient conditions at the early and middle-late stages of growth in calcareous soil, compared to both the non-transgenic line and lines harboring either OsIRT1 promoter-refre1/372 or 35S promoter-OsIRO2 alone. RI lines also exhibited a 9-fold higher yield than the non-transgenic line. Moreover, we successfully produced Fe-deficiency-tolerant Tachisugata rice, which is a high-biomass variety used as fodder. Collectively, our results demonstrate that combined enhancement of two Fe uptake systems in rice is highly effective in conferring tolerance to low Fe availability in calcareous soil.

TPS46, a Rice Terpene Synthase Conferring Natural Resistance to Bird Cherry-Oat Aphid, Rhopalosiphum padi (Linnaeus).

Plant terpene synthases (TPSs) are key enzymes responsible for terpene biosynthesis, and can play important roles in defense against herbivore attack. In rice, the protein sequence of TPS46 was most closely related to maize TPS10. However, unlike maize tps10, tps46 was also constitutively expressed in rice even in the absence of herbivore attack. Potential roles or constitutive emissions of specific volatiles may due to the constitutive expressions of tps46 in rice. Therefore, in the present study, RNA interference (Ri) and overexpression (Oe) rice lines were generated to investigate the potential function of TPS46 in Oryza sativa sp. japonica. Interestingly, the rice plants become more susceptible to Rhopalosiphum padi when expression of tps46 was silenced compared with Wt in greenhouse conditions. Artificial infestation bioassays further confirmed that Ri rice lines were susceptible to R. padi, whereas Oe rice lines were repellent to R. padi. Based on GC-MS and ToF-MS analysis, a total of eight volatile products catalyzed by TPS46 in rice were identified. Among them, only limonene and Eβf could be detected in all the Ri, Oe, and Wt lines, whereas other six volatiles were only found in the blend of volatiles from Oe lines. Moreover, the amount of constitutive limonene and Eβf in the Ri lines was significantly lower than in Wt lines, while the amounts of these two volatiles in the Oe line were obviously higher than in control rice. Our data suggested that the constitutive emissions of Eβf and limonene regulated by the constitutive expression of tps46 may play a crucial role in rice defense against R. padi. Consequently, tps46 could be a potential target gene to be employed for improving the resistance of plants to aphids.

Research on the thermal shifts and electron–phonon coupling parameters of Ri lines for Cr3+ ions in LaGaO3 and LaAlO3 crystals

The thermal shifts of Ri lines for Cr3+ ions in LaGaO3 and LaAlO3 crystals are investigated by an expression including both the static contribution due to thermal expansion of lattice and the vibrational contribution due to electron–phonon interaction. The results show that the relative importance of static contributions (characterized by the ratio between the static contribution and vibrational one) is larger than 20% for these systems. So the assumption in the previous papers that the static contributions are very small and can be neglected is doubtful. The true electron–phonon coupling coefficients α' obtained through the above expression are increased by about 30% and 40%, respectively, for LaGaO3:Cr3+ and LaAlO3:Cr3+ in comparison with the corresponding parameters α acquired previously by taking account of only the vibrational contribution. So, in the reasonable and precise studies of the thermal shifts and the electron–phonon parameters for a spectral line in crystals, both the static and vibrational contributions should be contained.

Performance of LS97-1610בSpencer’ soybean recombinant inbred line population segregating for resistance toFusarium virguliforme

Clark, W. D., Reyes-Valdes, M. H., Bond, J. and Kantartzi, S. K. 2013. Performance of LS97-1610בSpencer’ soybean recombinant inbred line population segregating for resistance to Fusarium virguliforme . Can. J. Plant Sci. 93: 1179–1185. Sudden death syndrome (SDS) is a devastating disease in soybean which is caused by Fusarium virguliforme. Sudden death syndrome resistance is a quantitative trait; therefore, development of resistant varieties requires understanding of complex genetics and environmental effect. In this study, we aimed to characterize 94 F5:8recombinant inbred (RI) lines derived from a cross between a resistant line for SDS, LS97-1610 and a susceptible cultivar, ‘Spencer’ and to identify sources of resistance. The RI lines were evaluated for their relative resistance (RR) to SDS along with the parents in four different environments; two years (2009 and 2010) and two locations (Carbondale and Valmeyer, IL). Analysis of variance revealed significant interactions between genotype and environment in each year of experimentation and in 2-yr combined data. Broad-sense heritability was estimated by ANOVA results, and it was moderate (61%). These results indicate that SDS resistance is partially unpredictable due to environmental influence. Ten RI were identified to have a RR with no significant differences from LS97-1610 and two of them had a RR lower than resistant parent. These stable RI across environments were used for further evaluation in breeding programs. Additionally, data obtained from field evaluation can be used in combination with molecular data to study the effect of quantitative trait loci (QTL) with the environment. The identification of common QTL across environments with consistent expression is the ultimate goal of every marker-assisted selection program.

IDENTIFICATION OF A MAJOR QUANTITATIVE TRAIT LOCUS CONFERRING RICE BLAST RESISTANCE USING RECOMBINANT INBRED LINES

Blast disease caused by Pyricularia oryzae is one of the limiting&lt;br /&gt;factors for rice production world wide. The use of resistant&lt;br /&gt;varieties for managing blast disease is considered as the most&lt;br /&gt;eco-friendly approaches. However, their resistances may be&lt;br /&gt;broken down within a few years due to the appearance of new&lt;br /&gt;virulent blast races in the field. The objective of the present&lt;br /&gt;study was to identify the quantitative trait locus (QTL) conferring&lt;br /&gt;resistance to blast disease using 126 recombinant inbred&lt;br /&gt;(RI) lines originated from a crossing of a durably resistant upland&lt;br /&gt;rice genotype (Laka) and a highly susceptible rice accession&lt;br /&gt;cultivar (Kencana Bali). The RI population was developed&lt;br /&gt;through a single seed descent method from 1997 to 2004.&lt;br /&gt;Resistance of the RI lines was evaluated for blast in an endemic&lt;br /&gt;area of Sukabumi, West Java, in 2005. Disease intensity of the&lt;br /&gt;blast was examined following the standard evaluation system&lt;br /&gt;developed by the International Rice Research Institute (IRRI).&lt;br /&gt;At the same year the RI lines were analyzed with 134 DNA&lt;br /&gt;markers. Results of the study showed that one major QTL was&lt;br /&gt;found to be associated with blast resistance, and this QTL was&lt;br /&gt;located near RM2136 marker on the long arm of chromosome&lt;br /&gt;11. This QTL explained 87% of the phenotypic variation with&lt;br /&gt;37% additive effect. The map position of this QTL differed&lt;br /&gt;from that of a partial resistant gene, Pi34, identified previously&lt;br /&gt;on chromosome 11 in the Japanese durably resistant variety,&lt;br /&gt;Chubu 32. The QTL, however, was almost at the same position&lt;br /&gt;as that of the multiple allele-resistant gene, Pik. Therefore, an&lt;br /&gt;allelic test should be conducted to clarify the allelic relationship&lt;br /&gt;between QTL identified in this study and the Pik. The RI lines&lt;br /&gt;are the permanent segregating population that could be very&lt;br /&gt;useful for analysing phenotypic variations of important agronomic&lt;br /&gt;traits possibly owned by the RI lines. The major QTL&lt;br /&gt;identified in this study could be used as a genetic resource in&lt;br /&gt;improvement of rice varieties for blast resistance in Indonesia

Transcriptional Activation of OsDERF1 in OsERF3 and OsAP2-39 Negatively Modulates Ethylene Synthesis and Drought Tolerance in Rice

The phytohormone ethylene is a key signaling molecule that regulates a variety of developmental processes and stress responses in plants. Transcriptional modulation is a pivotal process controlling ethylene synthesis, which further triggers the expression of stress-related genes and plant adaptation to stresses; however, it is unclear how this process is transcriptionally modulated in rice. In the present research, we report the transcriptional regulation of a novel rice ethylene response factor (ERF) in ethylene synthesis and drought tolerance. Through analysis of transcriptional data, one of the drought-responsive ERF genes, OsDERF1, was identified for its activation in response to drought, ethylene and abscisic acid. Transgenic plants overexpressing OsDERF1 (OE) led to reduced tolerance to drought stress in rice at seedling stage, while knockdown of OsDERF1 (RI) expression conferred enhanced tolerance at seedling and tillering stages. This regulation was supported by negative modulation in osmotic adjustment response. To elucidate the molecular basis of drought tolerance, we identified the target genes of OsDERF1 using the Affymetrix GeneChip, including the activation of cluster stress-related negative regulators such as ERF repressors. Biochemical and molecular approaches showed that OsDERF1 at least directly interacted with the GCC box in the promoters of ERF repressors OsERF3 and OsAP2-39. Further investigations showed that OE seedlings had reduced expression (while RI lines showed enhanced expression) of ethylene synthesis genes, thereby resulting in changes in ethylene production. Moreover, overexpression of OsERF3/OsAP2-39 suppressed ethylene synthesis. In addition, application of ACC recovered the drought-sensitive phenotype in the lines overexpressing OsERF3, showing that ethylene production contributed to drought response in rice. Thus our data reveal that a novel ERF transcriptional cascade modulates drought response through controlling the ethylene synthesis, deepening our understanding of the regulation of ERF proteins in ethylene related drought response.