籽粒镉低积累水稻地上部镉高积累遗传特性分析

农田土壤镉（Cd）污染日益严重,导致稻米Cd含量超标事件频繁出现,使粮食安全问题备受关注。因此,合理利用Cd污染农田、降低水稻籽粒Cd含量成为亟待解决的问题。籽粒Cd低积累水稻雅恢2816的地上部具有较强的Cd积累能力,研究旨在弄清其地上部Cd积累能力的遗传稳定性,进一步揭示控制该性状的遗传基础,为利用分子标记辅助选育地上部Cd富集能力强、籽粒Cd安全的水稻提供途径。以水稻雅恢2816和3个不同品种水稻分别组配获得的F₁为研究对象,分析地上部Cd积累相关性状的杂种优势。进一步以优势组合C268A/雅恢2816构建F₂作图群体,对地上部Cd积累相关性状进行QTL定位分析。结果表明：（1） F₁地上部Cd积累相关性状杂种优势明显,遗传稳定性强。地上部Cd积累相关性状属数量性状,F₂中/超亲分离现象明显。（2）在第4、6号染色体上共挖掘到1个控制水稻地上部生物量和3个控制地上部Cd积累量的QTL位点,分别为qSB-6、qSCdA-4、qSCdA-6-1和qSCdA-6-2,表型贡献率为10.6%—14.4%,且增效等位基因均来自雅恢2816。（3）地上部Cd积累量与地上部生物量、Cd含量,根、糙米的生物量、Cd积累量,根-地上部转移系数均呈极显著正相关,与地上部-籽粒转移系数呈极显著负相关,存在4个QTL集簇区Cl-4-1、Cl-6-1、Cl-6-2和Cl-6-3。（4）区间marker 04171-marker 04197控制着地上部生物量和Cd积累量,与控制糙米Cd含量的QTL不重叠。研究表明：籽粒Cd低积累水稻雅恢2816携带控制地上部Cd高积累的等位基因,可在世代间稳定遗传,QTL位点qSCdA-4、qSCdA-6-1、qSCdA-6-2是控制该性状的重要遗传基础,可为分子标记辅助选育地上部Cd高积累、籽粒Cd低积累水稻提供理论依据。     

Regulation of ammonium-induced proline accumulation in detached rice leaves

Accumulation of proline in response to NH₄Cl was studied indetached leaves of rice (Oryza sativa cv. Taichung Native1). Increasing concentrations of NH₄Cl from 50 to 200mMprogressively increased proline content and this was correlated with theincrease in ammonium content. Proline accumulation induced by NH₄Clwas related to proteolysis, an increase in ornithine--aminotransferaseactivity, a decrease in proline dehydrogenase activity, and a decrease inproline utilisation and could not be explained by NH₄Cl-inducedmodification in ¹-pyrroline-5-carboxylate reductase activity.The content of glutamic acid was decreased by NH₄Cl, whereas theincrease in arginine and ornithine contents was found to be associated with theincrease in proline content in NH₄Cl-treated detached rice leaves.     

Involvement of polyamines in the post-anthesis development of inferior and superior spikelets in rice

Early-flowered superior spikelets usually exhibit a faster grain filling rate and heavier grain weight than late-flowered inferior spikelets in rice (Oryza sativa L.). But the intrinsic factors responsible for the variations between the two types of spikelets are unclear. This study investigated whether and how polyamines (PAs) are involved in regulating post-anthesis development of rice spikelets. Six rice genotypes differing in grain filling rate were field grown, and PA levels and activities of the enzymes involved in PA biosynthesis were measured in both superior and inferior spikelets. The results showed that superior spikelets exhibited higher levels of free spermidine (Spd) and free spermine (Spm) and higher activities of arginine decarboxylase (ADC, EC 4.1.1.19), S-adenosylmethionine decarboxylase (SAMDC, EC 4.1.1.50) and Spd synthase (EC 2.5.1.16) than inferior spikelets at the early endosperm cell division and grain filling stage. The maximum concentrations of free Spd and free Spm and the maximum activities of ADC, SAMDC and Spd synthase were significantly correlated with the maximum cell division and grain filling rates, maximum cell number and grain weight. Application of Spd and Spm to panicles resulted in significantly higher rates of endosperm cell division and grain filling in inferior spikelets along with the activities of sucrose synthase (EC 2.4.1.13), ADP glucose pyrophosphorylase (EC 2.7.7.27) and soluble starch synthase (EC 2.4.1.21), suggesting that these PAs are involved in the sucrose-starch metabolic pathway. The results indicate that the poor development of inferior spikelets is attributed, at least partly, to the low PA level and its low biosynthetic activity.     

The appearance of polygalacturonase mRNA in tomatoes: one of a series of changes in gene expression during development and ripening

Tomato mRNA was extracted from individual fruits at different stages of development and ripening, translated in a rabbit reticulocyte lysate and the protein products analysed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis. The results indicate that there are at least two classes of mRNA under separate developmental control. One group of approximately six mRNAs is present during fruit growth and then declines at the mature-green stage. Another group of between four and eight mRNAs increases substantially in amount at the onset of ripening, after the start of enhanced ethylene synthesis by the fruit, and continues to accumulate as ripening progresses. Studies of protein synthesis in vivo show that several new proteins are synthesised by ripening fruits including the fruit-softening enzyme polygalacturonase. One of the ripening-related mRNAs is shown to code for polygalacturonase, by immunoprecipitation with serum from rabbits immunised against the purified tomato enzyme. Polygalacturonase mRNA is not detectable in green fruit but accumulates during ripening. It is proposed that the ripening-related mRNAs are the products of a group of genes that code for enzymes important in the ripening process.Abbreviation SDS sodium dodecyl sulfate     

The effect of NaCl on proline accumulation in rice leaves

The regulation of proline accumulation in detached leaves of rice(Oryza sativa cv. Taichung Native 1) was investigated.Increasing concentrations of NaCl from 50 to 200 mM progressivelyincreased proline content in detached rice leaves. NaCl induced prolineaccumulation was mainly due to the effect of both Na⁺ andCl^– ions. Proline accumulation caused by NaCl was related toprotein proteolysis, an increase in ornithine--aminotransferaseactivity,a decrease in proline dehydrogenase activity, a decrease in prolineutilisation,and an increase in the content of the precursors of proline biosynthesis,ornithine and arginine. Results also show that proline accumulation caused byNaCl was associated with ammonium ion accumulation.     

Iron nutrition affects cadmium accumulation and toxicity in rice plants

The effect of iron (Fe) nutrition on cadmium (Cd) toxicity and accumulation in rice plants was studied using a hydroponic system. The inhibitory effect of Cd on plant growth and chlorophyll content (SPAD value) was dependent on Fe level and the genotype. Malondialdehyde (MDA) content in leaves and roots was not much affected by an increased Cd stress at 0.171 mg l⁻¹ Fe, but it showed a rapid increase when the plants were exposed to moderate (1.89 mg l⁻¹) and high (16.8 mg l⁻¹) Fe levels. High Fe nutrition caused a marked reduction in Cd content in both leaves and roots. Fe content in plants was lower at high Cd (5.0 μM) stress than at low Cd (<1.0 μM) stress. Cd stress increased both superoxide dismutase (SOD) and peroxidase (POD) activities at low and moderate Fe levels. However, with high Fe level, it increased the POD activity, but reduced the SOD activity. Our results substantiate the hypothesis that cell membrane-bound iron transporter (carrier) involved in high-affinity iron transport systems can also transport Cd, and both these ions may compete for this common carrier. The study further showed that there were significant correlations between MDA and Fe contents in leaves and roots of rice plants. It is suggested that the occurrence of oxidative stress in plants exposed to Cd stress is mediated by Fe nutrition. The present results also show that Cd stress affects the uptake of Cu and Zn.     

Enigmatic double-stranded RNA in Japonica rice

We have found a linear, 16 kb, double-stranded RNA (dsRNA) in symptomless Japonica rice (Oryza sativa L.) that is not found in Indica rice (Oryza sativa L.). The dsRNA was detected in every tissue and at every developmental stage, and its copy number was approximately constant (about 20 copies/cell). Double-stranded RNA was also detected in two strains of Oryza rufipogon (an ancestor of O. sativa). Hybridization experiments indicated that the dsRNA of O. rufipogon was homologous but not identical to that of O. sativa. The sequence of about 13.2 kb of the dsRNA was determined and two open reading frames (ORFs) were found. The larger ORF (ORF B) was more than 12 351 nucleotides long and encoded more than 4 117 amino acid residues.     

A field study of pollen-mediated gene flow from Mediterranean GM rice to conventional rice and the red rice weed

The objective of this study was to assess the frequency of pollen-mediated gene flow from a transgenic rice line, harbouring the gusA and the bar genes encoding respectively, -glucuronidase and phosphinothricin acetyl transferase as markers, to the red rice weed and conventional rice in the Spanish japonica cultivar Senia. A circular field trial design was set up to investigate the influence of the wind on the frequency of pollination of red rice and conventional rice recipient plants with the transgenic pollen. Frequencies of gene flow based on detection of herbicide resistant, GUS positive seedlings among seed progenies of recipient plants averaged over all wind directions were 0.036 ± 0.006% and 0.086 ± 0.007 for red rice and conventional rice, respectively. However, for both red rice and conventional rice, a clear asymmetric distribution was observed with pollination frequency favoured in plants placed under the local prevailing winds. Southern analyses confirmed the hemizygous status and the origin of the transgenes in progenies of surviving, GUS positive plants. Gene flow detected in conventional rice planted at 1, 2, 5 and 10 m distance revealed a clear decrease with increasing distance which was less dramatic under the prevailing wind direction. Consequences of these findings for containment of gene flow from transgenic rice crops to the red rice weed are discussed. The precise determination of the local wind conditions at flowering time and pollination day time appear to be of primary importance for setting up suitable isolation distances.     

Influence of aluminum in nutrient solutions on chemical composition in upland rice cultivars

Summary Aluminum toxicity is an important growth limiting factor for upland rice production on oxisols of cerrado region in Brazil. Data related to the effect of Al on uptake of nutrients for rice crop are limited. The effect of five Al concentrations (0, 10, 20, 40 and 60 ppm) in culture solution on the chemical composition of 30 upland rice cultivars was studied.Aluminum concentration and content in plant tissues were increased with higher levels of Al in all cultivar. In the roots Al content was higher as compared with the tops. Critical toxic level of Al in the tops of 21 days old plants varied from 100 to 417 ppm depending on the cultivars. Rice cultivars responded differently to Al treatments with respect to nutrients uptake. Increased Al concentrations in the solution exerted an inhibiting effect on the concentrations and contents of N, P, K, Ca, Mg, S, Na, Zn, Fe, Mn, B and Cu. Thus the inhibition was more effective for macronutrients in the plant tops in following order: Mg>Ca>P>K>N>S>Na. Whereas for micronutrients it was in the order of Mn>Zn>Fe>Cu>B. Morphological, physiological and biochemical effects of Al, toxicity responsible for the reduction in plant nutrient uptake, are discussed.     

Changes in putrescine accumulation induced by agents affecting cytosolic pH in detached rice leaves

Effects of compounds that influenced cytosolic pH on the level of putrescine in detached rice leaves were examined. Permeant weak acids, isobutyric acid and propionic acid, increased the level of putrescine in detached rice leaves. Procaine and trisodium citrate, known to be permeant weak bases, on the other hand, decreased the level of putrescine. It seems possible that the level of putrescine in detached rice leaves is regulated by the cytosolic pH.     

Response of irrigated dry-seeded rice to nitrogen and phosphorus in a semi-arid environment

Summary Under semi-arid conditions, three field experiments were conducted at Gezira Research Station to determine response of irrigated dry-seeded rice (Oryza sativa L. var IR 2053-206-1-3-6) to addition of nitrogen and phosphorus fertilizers. The experimental treatments included the factorial combinations of seven levels of nitrogen applied as urea and four levels of phosphorus applied as super phosphate. Plant growth and grain yield were significantly and progressively increased with the rise in the levels of added nitrogen and phosphorus. However, response to added phosphorus was restricted by the applied level of nitrogen. The responses of grain yield to nitrogen and phosphorus levels are given by quadratic regression equations. Without addition of nitrogen or phosphorus grain yield averaged 1.52 t/ha compared to 6.07 t/ha with addition of the optimal levels (160 kg N plus 35 kg P/ha). The high potential for rice production in semi-arid environment is evidently restricted by addition of relatively high rates of nitrogen and phosphorus.     

Expression of functional proteins of cDNA encoding rice nucleoside diphosphate kinase (NDK) in Escherichia coli and organ-related alteration of NDK activities during rice seed germination (Oryza sativa L.)

The GST (glutathione S-transferase)-NDK (nucleoside diphosphate kinase) fusion protein was expressed in Escherichia coli. The GST-NDK protein was capable of transferring -phosphate from ATP to nucleoside diphosphates such as GDP, CDP, TDP and UDP. Western blot analysis using anti-NDK antibody indicated that NDK in endosperm gradually decreased during 36 h of imbibition. On the contrary, NDK in embryo increased during the same period. NDK activities in both tissues were in accord with these observations. Whereas the NDK protein in roots of rice seedlings during 7 days of imbibition remained constant, in shoots it declined after 5 days of imbibition. Thus, NDK may play a significant role in the cellular event modulated by adenylate energy charge level.     

Structural changes in the plastid DNA of rice (Oryza sativa L.) during tissue culture

To investigate the rearrangement of the plastid genome during tissue culture, DNA from rice callus lines, which had been derived individually from single protoplasts isolated from seed or pollen callus (protoclones), was analyzed by Southern hybridization with rice chloroplast DNA (ctDNA) clones as probes. Among 44 long-term cultured protoclones, maintained for 4, 8 or 11 years, 28 contained plastid DNA (ptDNA) from which portions had been deleted. The ptDNA of all protoclones that had been maintained for 11 years had a deletion that covered a large region of the plastid genome. The deletions could be classified into 15 types from their respective sizes and positions. By contrast, no deletions were found in the ptDNA of 38 protoclones that had been maintained for only 1 month. These results indicate that long-term culture causes deletions in the plastid genome. Detailed hybridization experiments revealed that plastid genomes with deletions in several protoclones were organized as head-to-head or tail-to-tail structures. Furthermore, ptDNAs retained during long-term culture all had a common terminus at one end, where extensive rearrangement is known to have occurred during the speciation of rice and tobacco. Morphological analysis revealed the accumulation of starch granules in plastids and amyloplasts in protoclones in which the plastid genome had undergone deletion. Our observations indicated that novel structural changes in the plastid genome and morphological changes in the plastid had occurred in rice cells during long-term tissue culture. Moreover, the morphological changes in plastids were associated with deletions in the plastid genome.     

Hormones in the grains in relation to sink strength and postanthesis development of spikelets in rice

Inferior spikelets usually exhibit a slower grain filling rate and lower grain weight than superior spikelets in a rice (Oryza sativa L.) panicle. This study investigated whether the variations in grain filling between the two kinds of spikelets were attributed to their sink strength and whether the sink strength was regulated by the hormonal levels in the grains. Using two field-grown rice genotypes, the division rate of endosperm cells, hormonal levels in the grains, and grain weight of both superior and inferior spikelets were determined during the grain filling period. The results showed that superior spikelets had dominance over inferior spikelets in endosperm cell division rate and cell number, grain filling and grain weight. Changes in zeatin (Z) and zeatin riboside (ZR) contents paralleled and were very significantly correlated with the cell division rate and cell number. Cell division rate and the content of indole-3-acetic acid (IAA) in the grains were also significantly correlated. Gibberellin (GAs; GA₁+ GA₄) content of the grains was high but ABA levels were low at the early grain filling stage. ABA increased substantially during the linear phase of grain growth and was very significantly correlated with grain dry weight during this period. Application of kinetin at 2 through 6 days post anthesis (DPA) significantly increased cell number, while spraying ABA at 11 through 15 DPA significantly increased the grain filling rate. The results suggest that differences in sink strength are responsible for variations in grain filling between superior and inferior spikelets. Both cytokinins and IAA in the grains may mediate cell division in rice endosperm at early grain filling stages, and therefore regulate the sink size of the grain, whereas ABA content correlates with sink activity during the linear period of grain growth.     

Activity of a maize ubiquitin promoter in transgenic rice

We have used the maize ubiquitin 1 promoter, first exon and first intron (UBI) for rice (Oryza sativa L. cv. Taipei 309) transformation experiments and studied its expression in transgenic calli and plants. UBI directed significantly higher levels of transient gene expression than other promoter/intron combinations used for rice transformation. We exploited these high levels of expression to identify stable transformants obtained from callus-derived protoplasts co-transfected with two chimeric genes. The genes consisted of UBI fused to the coding regions of the uidA and bar marker genes (UBI:GUS and UBI:BAR). UBI:GUS expression increased in response to thermal stress in both transfected protoplasts and transgenic rice calli. Histochemical localization of GUS activity revealed that UBI was most active in rapidly dividing cells. This promoter is expressed in many, but not all, rice tissues and undergoes important changes in activity during the development of transgenic rice plants.     

Ammonium in relation to proline accumulation in detached rice leaves

Ammonium accumulation in relation to prolineaccumulation in detached rice leaves under stressconditions was investigated. Ammonium accumulation indark-treated detached rice leaves preceded prolineaccumulation. Ammonium accumulation caused by waterstress coincided closely with proline accumulation indetached rice leaves. Exogenous NH₄Cl andmethionine sulfoximine (MSO), which caused anaccumulation of ammonium in detached rice leaves,increased proline content. It was found that prolinein NH₄Cl- or MSO-treated rice leaves is lessutilized than in water-treated rice leaves (controls). These results are in agreement with the observationthat a decrease in proline utilization contributes tothe accumulation of proline in dark-treated and waterstressed rice leaves. Although ammonium contentincreased in Cd- and Cu-treated rice leaves, theincrease in ammonium content was only observed afterthe increase in proline content.     

Induction of a polyubiquitin gene promoter by dehydration stresses in transformed rice cells

The expression of the maize polyubiquitin gene promoter UBI1 in rice cells has been used to study the involvement of ubiquitin in cell protection responses to dehydration caused by osmotic, saline or freezing stress. The effect of these stresses on UBI1 activity was investigated by the use of stably transformed rice calli (UBI1:GUS), as well as by transient expression experiments performed with cell lines with high or low tolerance to each type of stress. The theoretical analysis of the UBI1 promoter shows several putative stress-regulated boxes that could account for the stress-related UBI1 induction pattern described in this work. We suggest that the study of the differential UBI1 promoter-driven expression in rice cell lines with different level of tolerance to stress might be useful to elucidate complex signal transduction pathways in response to dehydration stresses in monocots.     

Pattern of dry-matter accumulation,and nitrogen concentration and uptake as influenced by levels and methods of nitrogen application in rainfed-upland rice

Summary Dry-matter accumulation, and concentration and uptake of nitrogen increased with increasing level of nitrogen at all the stages of crop growth. The differences in nitrogen concentration due to nitrogen levels were greatest at panicle initiation stage and started becoming narrower with the advancement in crop age. Split application of nitrogen with its heavier fractions at tillering and panicle initiation stages either through soil alone or soil+foliage (1/3+1/3+1/3) resulted in higher dry-matter accumulation, and higher nitrogen concentration and uptake than other methods. The crop, on an average, removed 61 kg N/ha. Plants accumulated nearly 15% of the total absorbed nitrogen, up to tillering, 50% up to panicle initiation and 85–90% up to heading. Proportionately lesser nitrogen uptake and dry-matter accumulation at post-heading stage is an indicative of a major constraint for production efficiency of rainfed-upland rice culture.     

Identification of a gibberellin-induced gene in deepwater rice using differential display of mRNA

Differential display of mRNA was employed to identify gibberellin (GA)-regulated genes in deepwater rice. One of the first differentially displayed products identified was shown to be ten-fold induced after start of GA treatment. The sequence of the clone shows complete amino acid identity with histone H3, and its increased mRNA level correlates with the onset of DNA synthesis. We also identified a gene whose expression pattern did not change over the course of treatment with GA and can be used as standard to correct for loading differences on northern blots.