Detection of changes in DNA methylation induced by low-energy ion implantation in Arabidopsis thaliana

This study evaluated changes in DNA methylation in Arabidopsis thaliana plants grown from seeds implanted with low-energy N(+) and Ar(+) ions. Methylation-sensitive amplified polymorphism (MSAP) testing revealed altered DNA methylation patterns after ion implantation at doses of 1 × 10(14) to 1 × 10(16) ions/cm(2). Comparison of the MSAP electrophoretic profiles revealed nine types of polymorphisms in ion-implanted seedlings relative to control seedlings, among which four represented methylation events, three represented demethylation events, and the methylation status of two was uncertain. The diversity of plant DNA methylation was increased by low-energy ion implantation. At the same time, total genomic DNA methylation levels at CCGG sites were unchanged by ion implantation. Moreover, a comparison of polymorphisms seen in N(+) ion-implanted, Ar(+) ion-implanted, and control DNA demonstrated that the species of incident ion influenced the resulting DNA methylation pattern. Sequencing of eight isolated fragments that showed different changing patterns in implanted plants allowed their mapping onto variable regions on one or more of the five Arabidopsis chromosomes; these segments included protein-coding genes, transposon and repeat DNA sequence. A further sodium bisulfite sequencing of three fragments also displayed alterations in methylation among either different types or doses of incident ions. Possible causes for the changes in methylation are discussed.     

Resistance in rice to Rice yellow mottle virus and evidence of non-seed transmission

Nine rice cultivars were evaluated under screenhouse conditions for resistance to Rice yellow mottle virus (RYMV) and possible seed transmission. Completely randomised design with three replications was used. In Experiment 1, the seedlings were inoculated with the virus at two weeks after planting. In Experiment 2, the seeds collected from Experiment 1 were dried for four weeks before planting. For each genotype, the seeds from healthy plants were planted as a control. Disease incidence and severity (scales 1–9), yield and yield components were recorded. Statistical analyses included Area Under the Disease Progress Curve (AUDPC) and independent t test. The cultivars FARO 37, FARO 52 and Gigante were highly resistant, whereas WAB189-B38HB was resistant. Paddy yield was highest (3.6 g) in FARO 37. There were no symptoms of virus disease in all the plants originating from the seeds of RYMV-infected plants. The differences between the seeds from infected and healthy plants for all the measured traits were not significant (p > 0.05). The number of days to seedling emergence was uniform (5.5 days) in all the cultivars. Plant height, number of tillers per plant, number of days to heading and paddy yield from the seeds of virus-infected plants varied from 54.8 to 68.4 cm, 17 to 21, 85.3 to 96 days and 2.7 to 4 g, respectively. Conversely, a range of 54.9–68.7 cm, 17–22, 83–95 days and 2.8–4.1 g was found in the seeds of healthy plants. Selection and cultivation of high-yielding, resistant and healthy seeds would enhance food security.     

Concentration of virus and ultrastructural changes in oats at various stages of barley yellow dwarf virus infection

Enzyme-linked immunosorbent assay was used to monitor the concentration of barley yellow dwarf virus (BYDV) in roots and leaves of oats inoculated at the 1 - 2 leaf stage and at the 4 - 5 leaf stage, respectively. Virus was detectable 20 h after inoculation in the roots and after 48 h in the leaves of plants inoculated at the 1 - 2 leaf stage. The virus concentration reached a plateau in the roots after 7–8 days, and was 3–4 times higher than in the leaves. In plants inoculated at the 4 - 5 leaf stage virus was detectable in roots and leaves after 3 and 5 days, respectively. The concentration reached a maximum after 10 days in the roots and after 18 days in the leaves; the concentration in the leaves was 2–3 times higher than in the roots. Virus was readily detectable in seeds from infected plants, both fresh and old dried seeds. However, seed transmission could not be demonstrated. Virus-like particles were first observed in phloem cells of roots 4 days after inoculation, but no ultrastructural changes were detected at this stage. After 5–6 days, disintegrated nuclei and virus-induced vesicles were observed in many cells and abnormal production of callose was found after 10 days. Necrotic phloem cells were observed from day 13, shortly after the appearance of external symptoms.     

Sexual modification of female spinach seeds (Spinacia oleracea L.) by irradiation with ion particles

The female seeds of a spinach plant (Spinacia orelacea L.) were exposed to He (12.5 MeV/n) and C (18.3 MeV/n) ions in order to investigate the effects of ion particles on sex expression. He ions did not affect germination rates or flowering at doses up to 50 Gy. C ions did not affect germination rates or flowering at doses up to 15 Gy, but a dose of 25 Gy resulted in many plants with morphological aberrations. When unexposed female plants were grown without cross-fertilization for 10 weeks after sowing, 5.6-14.3% of the plants produced anthers from female flowers. These sex-modified plants could self-pollinate and form seeds, which expressed only female organs. Conversely, gynomonoecious plants were induced from these female seeds by exposure to He ions (5-50 Gy) and C ions (5-25 Gy) without any difference in the rates of flowered progeny. Moreover, andromonoecious plants were induced from female seeds by exposure to He ions at 50 Gy. These results suggest that the sex of a spinach plant is expressed as a flexible phenotype, converging from female to gyno- and andromonoecy after exposure to ion particles.     

Genetically marked Rhizobium identifiable as inoculum strain in nodules of soybean plants grown in fields populated with Rhizobium japonicum

The fate of an inoculum strain of Rhizobium japonicum was studied using a genetically marked strain I-11O subline carrying resistance markers for azide, rifampin, and streptomycin (I-110 ARS). At the time of planting into a field populated with R. japonicum, seeds of soybean cultivars Kent and Peking were inoculated with varying cell densities of strain I-110 ARS. At various times during the growing season, surface-sterilized root nodules were examined for the presence of the inoculum strain by plating onto selective media. The recovery of the inoculum strain was unambiguous, varying, in the case of Kent cultivar, from about 5% with plants (sampled at 51 days) that had been inoculated with 3 X 10(8) cells per cm of row to about 20% with plants (sampled at 90 days) that had been inoculated with 3 X 10(9) cells per cm. The symbiotically incompatible interaction of Peking and strain 110 in Rhizobium-populated field soil was confirmed by the finding that at 60 days after planting, only one nodule in 360 sampled contained strain I-110 ARS. The use of genetically marked Rhizobium bacteria was found to provide for precise identification of the inoculum strain in nodules of field-grown soybeans.     

Genetically marked Rhizobium identifiable as inoculum strain in nodules of soybean plants grown in fields populated with Rhizobium japonicum.

The fate of an inoculum strain of Rhizobium japonicum was studied using a genetically marked strain I-11O subline carrying resistance markers for azide, rifampin, and streptomycin (I-110 ARS). At the time of planting into a field populated with R. japonicum, seeds of soybean cultivars Kent and Peking were inoculated with varying cell densities of strain I-110 ARS. At various times during the growing season, surface-sterilized root nodules were examined for the presence of the inoculum strain by plating onto selective media. The recovery of the inoculum strain was unambiguous, varying, in the case of Kent cultivar, from about 5% with plants (sampled at 51 days) that had been inoculated with 3 X 10(8) cells per cm of row to about 20% with plants (sampled at 90 days) that had been inoculated with 3 X 10(9) cells per cm. The symbiotically incompatible interaction of Peking and strain 110 in Rhizobium-populated field soil was confirmed by the finding that at 60 days after planting, only one nodule in 360 sampled contained strain I-110 ARS. The use of genetically marked Rhizobium bacteria was found to provide for precise identification of the inoculum strain in nodules of field-grown soybeans.     

Influence of cadmium and nickel on growth,net photosynthesis and carbohydrate distribution in rice plants

Six day old rice seedlings (Oryza sativa L. cv. Bahía) were grown for 5 or 10 days in a nutrient solution with either Cd (0.01, 0.1 mmol/l) or Ni (0.1, 0.5 mmol/l). Both Cd and Ni reduced the length of shoots and roots depending on the concentration and type of ion tested. On the other hand, the dry weight to fresh weight ratio was increased by heavy metal treatments, especially in the aerial part of 0.5 mmol/l Ni treated plants. The application of 0.1 mmol/l Cd and 0.5 mmol/l Ni to the seedlings produced an inhibition of the transport of carbohydrate reserves from the seeds from which plants were developing, to the rest of the plant. Net photosynthesis was also inhibited in treated plants. However, the total carbohydrate content in the shoots of these plants was higher than in controls. Thus, the starch, soluble sugars and sucrose content in the shoots of 0.5 mmol/l Ni treated plants was respectively up to 2.6, 2.8 and 4 times greater compared to controls. The distribution of assimilates between organs was also affected by the treatment: the carbohydrate content increased in the stem and second leaf but it was not affected or decreased in the root and third leaf. Although less evident, the effect of Cd on carbohydrate distribution and content was similar to that of Ni. The possible mechanisms involved in the abnormal carbohydrate accumulation and distribution are discussed.Abbreviations 0 DT plants at day zero of treatment - 5 DT five days treated plants - 10 DT ten days treated plants - DW dry weight - FW fresh weight     

Accumulation of ions during seed development under controlled saline conditions of two Suaeda salsa populations is related to their adaptation to saline environments

Controlled conditions were used to investigate the relationship between ion distribution in developing seeds of two Suaeda salsa populations and seed germination and seedling emergence. Seeds were harvested from S. salsa plants that had been treated with 1 or 400 mM NaCl for 122 (saline inland population) or 135 days (intertidal zone population) in a glasshouse. Germination and seedling emergence were evaluated under salinity. In both populations, more ions were accumulated in the pericarps of plants cultured in 400 mM NaCl than in 1 mM NaCl. Pericarps accumulated much higher ion concentrations in the intertidal zone population than in the saline inland population, while the opposite trend occurred for ion accumulation in the embryos. Seeds of plants from the intertidal zone population germinated more rapidly than those from plants of the saline inland population, regardless of the NaCl concentration during seed germination. However, seedling emergence under high salinity was lower with seeds from the intertidal zone population than with seeds from the saline inland population. In conclusion, S. salsa in the intertidal zone employs superior control of ion compartmentalization in the pericarps to tolerate salinity but requires a minimal level of ions in embryos to ensure seedling establishment in highly saline environments. This indicates that euhalophytes require salts during the mature seed stage to maintain seed viability and to ensure seedling emergence and population establishment.     

Some Physiological Parameters and Seed Lipid Composition of Transgenic Tobacco Plant

To see the effects of foreign gene introduction on the physiological performance and the quality and quantity of seed lipids, we studied transgenic tobacco plant as a model system, as tobacco seeds are oil seeds. Using Agrobacterium Ti plasmid based vectors, tobacco plants cv Petit Havana were transformed by NPT II gene as selectable marker. Transformed T₀ generation plants raised in tissue culture were transferred to pots and selfed. From the seeds, T₁ generation plants were grown in pots and their physiological performance was assessed. The transgenic plants showed slightly slower rates of germination and growth. Total chlorophyll content, chlorophyll a/b ratio and specific leaf weight, however, remained unchanged. The transgenic plants also had delayed flowering. However, total protein, lipid content and fatty acid composition of lipids of seeds in transgenic plants did not show appreciable difference from the seeds from control plants. Thus the physiological cost of transgenic plant for the extra genetic load was only marginal, if any.     

Hereditary variability of plants under the action of exogenous DNA

Summary Injection of exogenous barley donor DNA into grains of barley recipient plants at the milk maturity stage, with a specially designed syringe, led to the appearance of transformed plants. The transformation (in rare cases) was caused by the unsheared DNA since the DNA passing through the syringe needle remained relatively stable (10⁶ to 10⁷ daltons) as was confirmed by DNA sedimentation analysis.14 plants grown from seeds injected with highly polymeric DNA containing close to 30 per cent protein had transformed pollen grains. In the 2nd generation only 2 plants from the 8 studied preserved these changes. In the progeny of these two plants, i.e., in the 3rd seed generation after injection, 82.1 per cent of plants preserved the transformed characters. The next, 4th generation, preserved a transformed phenotype in 89.6 per cent of plants.It was also shown that reversion to a recipient-like state was not always constant. We found the reversion of transformed properties (i.e., normal starch and two-rowed spikes) in 40 per cent of the 4th generation descendants of one of the plants which had lost the phenotypical expression of these properties in the 3rd generation but had them in the 2nd generation.The study of the morphological properties of transformed plants showed that with respect to phenotypic expression some characters were changed towards the donor type, some remained as in the recipients and some were of the intermediate type.     

盐地碱蓬二型性种子及其幼苗对盐渍环境的适应性

研究了盐地碱蓬二型性种子中离子含量与刚萌发幼苗耐盐性之间的关系,以及盐分对砂培盐地碱蓬二型性种子的幼苗生长、离子含量及光合特性的影响.棕色种子中离子含量显著高于黑色种子.与对照相比,100和400 mmol/L NaCl对棕色种子幼苗伸长没有抑制作用,却显著抑制黑色种子幼苗的伸长.NaCl处理下棕色种子的幼苗地上部分干重和主茎一级分枝数比黑色种子幼苗高,但二型性种子的幼苗叶片中离子含量、叶绿素含量及光合放氧速率却没有明显差异.上述结果说明盐地碱蓬棕色种子较高的离子含量可能是棕色种子刚萌发幼苗耐盐性较强的重要原因.棕色种子幼苗较高的生物量可能与其较多的分枝数有关.二型种子的这些特征可能决定了其在群落建成中所起到的不同作用.     

Development of somatic embryos from cell suspension cultures of Norway spruce (Picea abies Karst.)

Cell suspension cultures were established from embryonal-suspensor masses derived from mature seeds. Transfer of cell masses on to a medium containing abscisic acid stimulated development of already established individual embryos. Somatic embryos developed shoots when supported by cheese cloth in liquid medium in Petri dishes. The percentage of well-developed roots remained low even though all embryos had root meristems. We have recovered an average of 25 plantlets from an initial PCV of ca 1 g fresh wt per 10 ml.     

低温锻炼对绵头雪莲花组织培养苗抗寒性及抗氧化酶活性的影响

高山植物绵头雪莲花(Saussurea laniceps HandMazz.)组织培养苗经2℃低温锻炼后抗寒性明显增强,SOD、CAT、POD活性明显提高,可溶性蛋白质、脯氨酸含量升高,而可溶性糖含量没有明显变化.脱锻炼期间可溶性蛋白质含量仍高于低温锻炼期间的水平,脱锻炼后抗氧化酶活性略有上升或保持稳定状态.这些变化是绵头雪莲花组织培养苗适应低温逆境的生理生化基础.     

建立低能离子束介导小麦转基因方法并获得转GUS基因植株

研究了注入离子种类、能量、剂量等参数对于低能离子束介导的遗传转化的影响,建立了适于小麦成熟胚转化的组培条件和筛选程序。以携带ＧＵＳ基因的质粒为供体,进行了报告基因转化研究。分子生物学证据表明ＧＵＳ基因已整合到小麦基因组中。３个小麦品种的抗性愈伤转化率分别为９．５％、１０．８％、１１．２％,再生植株转化率分别为１．４％、３．４％、１．７％,首次证明了离子束介导小麦遗传转化是可行的,为离子束介导小麦遗     

鹤望兰组织培养与工厂化快繁程序的研究

将材料接种于诱导愈伤组织手芽的培养基上,培养２个月后,胚芽外植体下出现白色颗粒状的愈伤组织,４个月后愈伤组织上出现小芽丛。将小芽丛转入不加植物激素的ＭＳ培养基上,芽的生长加快,２个月左右可长成３－６ｃｍ高的丛小植株。将小植株切下,插入根培养基中,一般３５ｄ左右基部突出很小的白色根尖。     

Plant regeneration from seedling explants of green bean (Phaseolus vulgaris L) via organogenesis

Green bean (Phaseolus vulgaris L.) plants were regenerated from 3-day old seedling explants via organogenesis. The explants contained a cotyledon and a small portion (2–3 mm) of embryonic axis split in half. Explants were cultured on a defined medium containing glutamine as the sole nitrogen source. A ring of meristematic tissue was produced at the base of the axillary bud located at the cotyledonary node. The meristematic tissue was produced only if the axillary bud was present together with the cotyledon in the explant. Buds and shoots developed from the meristematic ring. Selected shoots produced roots when excised from the cluster of buds and transferred to root induction medium. Rooted shoots (plantlets) grew well and produced viable seeds when grown in the greenhouse. Histological studies revealed the origin of buds from the peripheral layers of the meristematic ring.Production of buds and shoots was a continuous process, so that new shoots could be removed from the explant for plantlet production every 10–14 days. With the cultivar Dark Red Kidney, an average of 49 buds and 8 shoots were regenerated per explant by 30 days after culture initiation. Sixty-seven percent of the shoots produced roots, and 90–95% of the plantlets survived greenhouse acclimatization to produce healthy plants.     

Ecological significance of the aerial seed pool of a desert lignified annual,Blepharis sindica (Acanthaceae)

Reproductive traits of a lignified annual plant, Blepharis sindica were studied in relation to the formation of an 'aerial seed pool' on dead plants in an arid grassland in the Thar Desert of northwestern India. The dead plants remained standing on the soil surface and retained fruits for more than one year. Aerial seed pools developed about 6 cm above the ground. There were no seed pools on or below the ground surface. Only 5.7% of seeds died on dead plants because of insect predation or fungi infection during one year. Seed release was cued by rainfall, and a fraction of seeds on the aerial seed pools was released in each rainfall event. After 13 rainfall events during the monsoon season, 25% of seeds was still retained on the plants. Seed predation on the ground surface was intensive; all cones placed on the soil surface were removed within four days, and 97% of fruits were removed within 10 days. Fifty percent of seeds germinated within 3.5 h, and there were no differences in viability and time required for germination between first year seeds and older seeds. The results indicate that the aerial seed-holding on dead plants is an available way to avoid seed predation in harsh desert environments where seed predation is intense and favorable periods for growth are temporally limited and unpredictable.     

农杆菌介导的雪花莲凝集素基因转入玉米骨干自交系

以农杆菌AGL0介导,将雪花莲凝集素基因转入玉米骨干自交系齐319和掖515胚性愈伤组织细胞,从筛选后的抗性愈伤组织获得再生植株。农杆菌浓度和共培养时间均能显著影响侵染后玉米愈伤组织的抗性频率。在农杆菌浓度OD600 0．2～0．3,共培养时间3d时,侵染后玉米愈伤组织的抗性频率最高,平均约4％。对再生植株及其子代基因组DNA的PCR及Southern杂交分析表明雪花莲凝集素基因已经整合到玉米基因组中,并遗传给后代。在蚜虫人工接种试验中,转基因植株上蚜虫的繁殖力为非转基因对照植株上的50％,这表明转基因植株抗蚜性显著增强。     

Growth and certain chemical constituents of tobacco plants exposed to air ions

Controlled experiments were performed in Faraday cages on the effects of positive and negative air ions on flue-cured tobacco plants. Continuous exposures for 15 days to air ions showed no significant differences in any plant growth characteristic between the treated and control plants. Standard errors in the measurement of the growth parameters for ion exposed plants were, however, consistently higher than those of control plants. Spatial variation in concentration gradients of air ions produced by corona discharge might have contributed to masking of the relatively small effects of air ions on biological organisms observed in previous experiments in this laboratory. No significant difference was observed between the experimental and control plants in nicotine, total alkaloid, and reducing sugar contents. Total nitrogen content was slightly higher for treated than control plants.     

香荚兰的快速繁殖

香荚兰是生长在热带地区的攀援性兰科植物。原产墨西哥,现广泛栽植在热带多雨地区。它的果实(荚)发酵后,是一种天然食品香料(主香成份是香草醛)的来源。 香荚兰可以用种子繁殖,但从出芽到结实约需7—9年。因此,通常用扦插法繁殖。用长为30—50厘米的插条扦插,3—4年后即可开花结实。因所用的插条较长,国