Possible Metsulfuron Herbicide Detoxification by a Oryza sativa L. Glutathione S-transferase Enzyme

Abstract Rice (Oryza sativa L.) is one of the most important crops in the world, and it is considered the primary source of nutritional layout in developing countries in Asia. The glutathione S-transferases (GSTs) superfamily confers to rice protection against biotic and abiotic stress, and herbicide resistance. However, the three-dimensional structure of a GST Tau class, is unsolved. The objectives of this work were to develop a reliable comparative model for the s-transferase glutathione class Tau 4 from rice, and simulate docking interactions, against herbicides bentazon and metsulfuron. Results showed that the predicted model is reliable and has structural quality. Ramachandran plot set 91.9% of the residues in the most favored regions. All complexes showed negative binding energies values; and metsulfuron docked to the glutathione tripeptide, and it represents a possible insilico evidence of glutathione conjugation with this herbicide.

Transcriptome analysis of phosphorus stress responsiveness in the seedlings of Dongxiang wild rice (Oryza rufipogon Griff.)

Abstract Background: Low phosphorus availability is a major factor restricting rice growth. Dongxiang wild rice (Oryza rufipogon Griff.) has many useful genes lacking in cultivated rice, including stress resistance to phosphorus deficiency, cold, salt and drought, which is considered to be a precious germplasm resource for rice breeding. However, the molecular mechanism of regulation of phosphorus deficiency tolerance is not clear. Results: In this study, cDNA libraries were constructed from the leaf and root tissues of phosphorus stressed and untreated Dongxiang wild rice seedlings, and transcriptome sequencing was performed with the goal of elucidating the molecular mechanisms involved in phosphorus stress response. The results indicated that 1184 transcripts were differentially expressed in the leaves (323 up-regulated and 861 down-regulated) and 986 transcripts were differentially expressed in the roots (756 up-regulated and 230 down-regulated). 43 genes were up-regulated both in leaves and roots, 38 genes were up-regulated in roots but down-regulated in leaves, and only 2 genes were down-regulated in roots but up-regulated in leaves. Among these differentially expressed genes, the detection of many transcription factors and functional genes demonstrated that multiple regulatory pathways were involved in phosphorus deficiency tolerance. Meanwhile, the differentially expressed genes were also annotated with gene ontology terms and key pathways via functional classification and Kyoto Encyclopedia of Gene and Genomes pathway mapping, respectively. A set of the most important candidate genes was then identified by combining the differentially expressed genes found in the present study with previously identified phosphorus deficiency tolerance quantitative trait loci. Conclusion: The present work provides abundant genomic information for functional dissection of the phosphorus deficiency resistance of Dongxiang wild rice, which will be help to understand the biological regulatory mechanisms of phosphorus deficiency tolerance in Dongxiang wild rice.

Cytogenotoxicity of rice crop water after application of the tricyclazole fungicide

ABSTRACT Tricyclazole is currently one of the fungicides recommended for the treatment of diseases in irrigated rice. However, there is relatively little information on its cytotoxic and genotoxic potential. The objective of this study was to evaluate the cytotoxicity and genotoxicity of rice crop water after apllication of the tricyclazole fungicide through the Allium cepa L. test. The rice crop water samplings were collected before and 1, 15 and 30 days after application of the fungicide in rice plant shoots. The Allium cepa roots were placed in contact with the rice crop water to check for possible chromosomal abnormalities and mitotic index of the bioindicators meristematic cells. The data obtained by the Allium cepa test indicates that the application of the tricyclazole fungicide leads to an increase in the genotoxic activity in the rice crop water, through the appearance of chromosomal abnormalities, without, however, causing significant effects on the mitotic index. The major chromosomal alterations observed were anaphasic and telophasic bridges and laggard chromosomes.

Effect of phosphorus on arsenic uptake and metabolism in rice cultivars differing in phosphorus use efficiency and response

ABSTRACT A hydroponic experiment was carried out to investigate the effect of phosphorus (P) nutrition on arsenic (As) uptake and translocation within the seedlings of rice cultivars. The experiment occurred in three stages: I 5 days of acclimatization (nutritive solution); II 10 days under P (0.0 and 0.09 mM) and As (0.0 and 100 mM) treatments; III 5 days under recovery. The As exposure had significant effect reducing dry weights of shoots or roots, resulted in elevated concentrations of As in shoot tissues. BR-IRGA 409 showed the highest susceptibility to As in biomass production and root system parameters regardless the P level. This cultivar showed contrasting responses of As translocation to shoot tissue dependent on P levels, with the highest As concentration under low P and lowest under normal P levels. P nutrition was most striking on plants recovery for all cultivars under As exposure. Clearer separation of cultivars for phosphorus use efficiency (PUE) occurred at lower shoot P contents, that was, at higher levels of P deficiency stress. IRGA 424 showed higher PUE as compared to the others cultivars. Our results go some way to understanding the role of P nutrition in controlling the effects of As in rice shoots.

Saline-induced changes of epicuticular waxy layer on the Puccinellia tenuiflora and Oryza sativa leave surfaces

BACKGROUND: The epicuticular waxy layer of plant leaves enhances the extreme environmental stress tolerance. However, the relationship between waxy layer and saline tolerance was not established well. The epicuticular waxy layer of rice (Oryza sativa L.) was studied under the NaHCO3 stresses. In addition, strong saline tolerance Puccinellia tenuiflora was chosen for comparative studies. RESULTS: Scanning electron microscope (SEM) images showed that there were significant changes in waxy morphologies of the rice epicuticular surfaces, while no remarkable changes in those of P. tenuiflora epicuticular surfaces. The NaHCO3-induced morphological changes of the rice epicuticular surfaces appeared as enlarged silica cells, swollen corns-shapes and leaked salt columns under high stress. Energy dispersive X-ray (EDX) spectroscopic profiles supported that the changes were caused by significant increment and localization of [Na(+)] and [Cl(-)] in the shoot. Atomic absorption spectra showed that [Na(+)]shoot/[Na(+)]root for P. tenuiflora maintained stable as the saline stress increased, but that for rice increased significantly. CONCLUSION: In rice, NaHCO3 stress induced localization and accumulation of [Na(+)] and [Cl(-)] appeared as the enlarged silica cells (MSC), the swollen corns (S-C), and the leaked columns (C), while no significant changes in P. tenuiflora.

Efficient in vitro plant regeneration via leaf base segments of indica rice (Oryza sativa L.).

A reliable and reproducible protocol has been developed for high frequency plant regeneration from 4-5 mm long leaf base segments of 4 days old in vitro germinated seedlings of indica rice (Oryza sativa) cultivar Rasi. The effect of age of seedlings, position of leaf base segments and optimum concentration of 2,4-D on callus induction frequency was investigated with a future aim to use leaf bases for biolistic and Agrobacterium-mediated transformation experiments. Friable, nodular and white to pale yellow embryogenic callus cultures (206 mg fresh weight /explant) were obtained from the first basal segments of rice seedlings on Linsmaier and Skoog (LS) medium supplemented with 2,4-D (11.3 microM) and 3.0 microM thiamine-HCL. Plant regeneration was achieved after the transfer of 54 days old embryogenic callus cultures to Murashige and Skoog (MS) medium supplemented with BAP (2.2 microM) and NAA (0.27 microM). In vitro regenerated plants with multiple shoots and roots transferred to sterile soil in growth chamber and maintained in greenhouse exhibited normal growth and were phenotypically similar to plants germinated from seeds.

Effect of salinity stress on growth and carbohydrate metabolism in three rice (Oryza sativa L.) cultivars differing in salinity tolerance.

Rice seedlings cv. Khao Dawk Mali 105 (salt-sensitive), Luang Anan (moderately salt-tolerant) and Pokkali (salt-tolerant) were exposed to 0, 50, 100 and 150 mM NaCI for 9 d. Salinity stress caused reduction in leaf relative water contents in all cultivars. Shoot length of cv. Pokkali was least affected by salinity stress whereas increased root length in response to salinity stress was apparent in cvs. Khao Dawk Mali 105 and Luang Anan. Increased salinity level also caused reduction in fresh and dry weights in cvs. Khao Dawk Mali 105 and Luang Anan, but had no effect in cv. Pokkali except at 150 mM. Accumulation of total soluble sugars and sucrose in mature leaves were observed in cv. Khao Dawk Mali 105 exposed to high level of salinity whereas their concentrations in cvs. Luang Anan and Pokkali remained the same as control plants. Accumulation of sucrose in cv. Khao Dawk Mali 105 was suggested to be resulted from the alteration of photosynthate partitioning since the activities of sucrose phosphate synthase were not affected by salinity in this cultivar. On the contrary, salinity stress induced an accumulation of starch in cv. Pokkali. It is suggested that partitioning sugars into starch may involve in salinity tolerance by avoiding metabolic alterations.

Pollution level in distillery effluent and its phytotoxic effect on seed germination and early growth of maize and rice.

The effluent from a Lucknow- based distillery (Mohan Meakin Distillery) was analyzed for physico-chemical and biological parameters of pollution and concentration of potentially toxic heavy metals (Cd, Cr, Ni and Zn) and the effect of the distillery effluent, as such and on 50% dilution with tap water was studied on seed germination and seedling growth of maize (Zea mays L.) and rice (Oryza sativa L.). The effluent was wine red in colour and highly acidic (pH approximately 55) and possessed decaying alcoholic smell. The effluent contained high values of different pollution parameters, particularly total solids, 3450 mgl(-1) (soluble plus suspended solids), alkalinity 1500 mgl(-1), biological oxygen demand (BOD, 1649 mgl(-1)) and chemical oxygen demand (COD, 2036 mgl(-1)). It had very low values of dissolved oxygen (DO, 0.34 mgl(-1)). The heavy metals (Cd, Cr, Ni and Zn) content, particularly the nickel concentration (0.029 mg l(-1)) was high. Use of the distillery effluent, even on 1:1 dilution with tap water inhibited germination and early seedling growth of maize and rice. In both maize and rice, more so in the former germination % of seeds, length of radicle and plumule and the fresh and dry weight of the seedlings were significantly reduced. The emerging leaves of the seedlings also developed visible effects of toxicity some of which resembled the symptoms of nickel toxicity. Our observations suggest that the effluent, as discharged from the distillery carry a heavy load of pollutants. Its discharge into the river Gomti poses a potential threat to the aquatic life, perticularly during the summer months when the water flow in the river is drastically reduced. The distillery effluentis also harmful for irrigating crops grown along the drain carrying it.

Functional validation of a novel isoform of Na+/H+ antiporter from Pennisetum glaucum for enhancing salinity tolerance in rice.

Salt stress is an environmental factor that severely impairs plant growth and productivity. We have cloned a novel isoform of a vacuolar Na+/H+ antiporter from Pennisetum glaucum (PgNHX1) that contains 5 transmembrane domains in contrast to AtNHX1 and OsNHX1 which have 9 transmembrane domains. Recently we have shown that PgNHX1 could confer high level of salinity tolerance when overexpressed in Brassica juncea. Here,we report the functional validation of this antiporter in crop plant rice.Overexpression of PgNHX1 conferred high level of salinity tolerance in rice. Transgenic rice plants overexpressing PgNHX1 developed more extensive root system and completed their life cycle by setting flowers and seeds in the presence of 150 mM NaCl. Our data demonstrate the potential of PgNHX1 for imparting enhanced salt tolerance capabilities to salt-sensitive crop plants for growing in high saline areas.

Candidate genes for drought tolerance and improved productivity in rice (Oryza sativa L.).

Candidate genes are sequenced genes of known biological action involved in the development or physiology of a trait. Twenty-one putative candidate genes were designed after an exhaustive search in the public databases along with an elaborate literature survey for candidate gene products and/or regulatory sequences associated with enhanced drought resistance. The downloaded sequences were then used to design primers considering the flanking sequences as well. Polymerase chain reaction (PCR) performed on 10 diverse cultivars that involved Japonica, Indica and local accessions, revealed 12 polymorphic candidate genes. Seven polymorphic candidate genes were then utilized to genotype 148 individuals of CT9993 x IR62266 doubled haploid (DH) mapping population. The segregation data were tested for deviation from the expected Mendelian ratio (1:1) using a Chi-square test (less than 1%). Based on this, four candidate genes were assessed to be significant and the remaining three, as non-significant. All the significant candidate genes were biased towards CT9993, the female parent in the DH mapping population. Single-marker analysis strongly associated (less than 1%) them to different traits under both well-watered and low-moisture stress conditions. Two candidate genes, EXP15 and EXP13, were found to be associated with root number and silicon content in the stem respectively, under both well-watered and low-moisture stress conditions.

Effect of 2,4-D, hydric stress and light on indica rice (Oryza sativa) somatic embryogenesis

With the purpose of increasing the embryogenesis regeneration process in vitroplants obtained from somatic embryos of the indica rice variety CR-5272 (Oryza sativa L.), two independent experiments were performed. The first experiment consisted in the effect of combination of three concentrations of the gelling agent PhytagelTM (1.8, 2.4, and 3 gL-1) and four 2,4-D concentrations (2.26, 4.52, 6.78, and 9.05 m M) on the induction and subsequent regeneration of embryogenic calli. On the second experiment, the pre-regeneration phase was modified; calli were subjected to darkness or diffuse light conditions for one, two, and three weeks. In embryogenesis induction, 35% calligenesis was obtained using the MS culture medium supplemented with 6.78 m M of 2,4-D and 2.4 gL-1 PhytagelTM , whereas on the control treatment (MS medium supplemented with 9.05 m M of 2,4-D and 3 gL-1 PhytagelTM ) 24% calligenesis was obtained. In addition, regeneration percentages were improved (22% and 16% for calli induced with the above treatments, respectively). Furthermore, in light exposure experiments, the best result was obtained by exposing the embryogenic calli to darkness for one week in pre-regeneration, followed by direct light exposure during the regeneration phase

Interference of three weed extracts on uptake of nutrient in three different varieties of paddy through radio tracer techniques.

Interference of three dominant weed extracts viz., Ageratum conyzoides L., Melilotus indica All. and Parthenium hysterophorus L. were examined on seed germination, seedling growth, and nutrient uptake (32P and 65Zn) in three different varieties (PD-10, PD-12 and PB) of paddy (Oryza sativa L.). Among the three different varieties irrespective of weed extracts, PD-10 and PD-12 were resistant and PB was susceptible in terms of seed germination, radicle length and plumule dry weight; and PD-12 and PB were resistant and susceptible, respectively, in terms of plumule length and total seedling dry weight. A. conyzoides caused maximum reduction in seed germination and M. indica in seedling growth in different varieties of paddy. The weed extracts interfered in uptake of both 32P and 65Zn and there was a gradual decrease in uptake of both nutrients with increasing concentration of extracts in both root and shoot. The uptake of 32P and 65Zn was more inhibitory with the extracts of A. conyzoides and M. indica, respectively in different varieties. The inhibition in seed germination, seedling growth and nutrient uptake may be due to the presence of phenolics and other secondary metabolities. The phenolics such as gallic, vanillic, protocatechuic and p-hydroxybenzoic acids were identified from these weed extracts.

RAPD analysis of herbicide-resistant Brasilian rice lines produced via mutagenesis

Over the last two decades, mutational techniques have become one of the most important tools available to progressive rice- breeding programs. In a mutation-breeding program initiated in 1999 at the Instituto Agronômico of Campinas, SP, Brazil, a rice line, IAC103, was selected for mutational studies with gamma radiation and ethyl methyl sulfonate mutagenesis, with the aim of developing a herbicide-resistant crop. After mutagenesis, surviving plants were exposed to glufosinate to check for herbicide resistance, which was examined up to the second generation. A detailed RAPD analysis was made of the resistant plants. Eighty Operon technology primers were tested and 10 were selected for a detailed study of RAPD markers that could tag herbicide resistance genes. Resistant and susceptible lines produced variation in the RAPD patterns and certain bands were found only in certain lines. These results suggest genetic ligation that will be confirmed through a genetic segregation study

Influence of cell wall degrading enzymes on colonization of N2 fixing bacterium, Azorhizobium caulinodans in rice.

In rice, nodule like structures were formed by inoculation of A. caulinodans combined with growth regulators and enzymes. Among the treatments, combination of cell wall degrading enzyme mixture and NAA with A. caulinodans induced more number of paranodules in rice. Total nitrogen content also increased in treated plants compared to uninoculated control.

Development and characterization of salt tolerant somaclones in rice cultivar pokkali.

Primary regenerants (1190) of a tall traditional salt tolerant rice cultivar pokkali were produced through in vitro culture from mature seed derived calli of fourth subculture. Out of 35000 SC2 regenerants, 26 promising lines with superior agronomic traits were chosen initially for evaluation. SC3 and SC4 generations were stringently evaluated under hydroponics with excess salt stress as well as under field conditions across two growing seasons in Bay Islands. A set of 10 promising somaclones was further evaluated at SC5 and SC6 of which BTS 2, BTS 13, BTS 18 and BTS 24 were found promising. In SC7 and SC8 yield trials in research farm, BTS 24 was found to produce a mean yield of 36.3 and 45.9 q ha-1 under saline and normal soil conditions, respectively. Somaclones varied significantly from the parent with respect to yield and yield attributes. Grain quality and biochemical parameters of all elite somaclones were different from the parent. However, somaclones did not deviate much from their parent in respect of disease and insect pest resistance pattern.

Enhancement of polygalacturonase activity during auxin induced para nodulation and endorhizosphere colonization of Azospirillum in rice roots.

Effect of different auxins, namely, 2,4-dichlorophenoxyacetic acid (2,4-D), naphthalene acetic acid (NAA) and indole acetic acid (IAA) and Azospirillum brasilense bioinoculation on the enhancement of polygalacturonase (PG) activity in rice roots during para nodulation and endorhizosphere colonization of Azospirillum was studied under in vitro condition. It was observed that Azospirillum bioinoculation could augment PG activity of rice roots to a lesser extent without any root morphogenesis whereas auxin application together with Azospirillum bioinoculation enhanced PG activity of rice roots to a higher level which resulted in better root morphogenesis (para nodule) and endorhizosphere colonisation of A. brasilense. Among the three auxins tested, 2,4-D, even at lower concentration (0.5 ppm) enhanced the rice root PG activity, root morphogenesis and endorhizosphere colonization of Azospirillum while it was 2.0 ppm with NAA and variable with IAA. It is concluded that there is a positive correlation existing among PG activity, degree of root morphogenesis and endorhizosphere colonization of Azospirillum brasilense in rice roots and the degree of correlation is determined by the chemical composition, concentration and mode of action of the auxin utilised.

Identification and characterization of some PR-proteins induced by kitazin and Rhizoctonia solani causing sheath blight of rice.

Significant reduction (68.38%) in sheath blight disease of rice was noticed when foliar spray of a systemic fungicide, kitazin (480 micrograms mg-1), was applied twice at an interval of 2 days before inoculation. SDS-PAGE analysis of proteins of Rhizoctonia-infected rice leaf sheaths revealed the presence of 16 proteins ranging from 20 to 90 kDa (approx.). Six were identified as constitutive defense proteins (increased after infection), 6 as secondary defense proteins (formed de novo) and the rest 4 appeared non-defense proteins. Non-inoculated kitazin-treated leaf sheaths showed 15 proteins of which 5 were constitutive and 4 secondary defense proteins (both are PR-proteins). Among the PR-proteins, five beta-1,3-glucanases and one chitinase was identified and characterized. One rice chitinase (MW 20 kDa) and 2 glucanases (60 & 69 kDa) showed serological relationships with tobacco chitinase (32 kDa) and tobacco glucanase (33 kDa) respectively. The implications of results have been discussed in relation to biotic and abiotic induction of PR-proteins in rice.