Plant regeneration from protoplasts isolated from suspension cultures of leek (Allium ampeloprasum L.)

A procedure is described for the regeneration of plants from protoplasts of tetraploid leek (Allium ampeloprasum L.), 2n = 4x =32. Regeneration-competent protoplasts could only be obtained from an embryogenic suspension culture that was initiated with friable, embryogenic callus derived from immature embryos. The generally low plating efficiency could be increased by embedding the protoplasts in Ca-alginate, compared to culturing the protoplasts in liquid or agarose-solidified medium. A minimum plating density of 2 × 10⁵ pps/ml was required to obtain microcalli. Upon transfer of the protoplast-derived calli on agarose-solidified BDS medium, morphologically different callus types proliferated. After transfer to regeneration medium, compact or friable calli with an embryogenic appearance produced somatic embryos and plantlets at a frequency of up to 80%. Calli that had been classified as heterogeneous also regenerated shoots, but mainly via organogenesis, at a frequency of 46%. After transfer of shoots to half strength MS medium, healthy, well-rooted plants were obtained, that were successfully transferred to soil. All plants contained the tetraploid DNA level.     

黄河三角洲海岸带湿地柽柳在干旱年份的水分利用策略

以黄河三角洲海岸带贝壳堤湿地灌木群落主要建群种柽柳为对象,利用稳定同位素技术测定柽柳木质部和潜在水源δ¹⁸O值的时空变化,采用IsoSource模型计算潜在水源对柽柳的贡献比例,研究海岸带不同生境中柽柳对不同水分条件的适应机制.结果表明: 在降水较少的干旱年份,相对于不稳定的降水,柽柳倾向于利用稳定的土壤水和浅层地下水,但是在不同微地形生境下柽柳的水分利用策略有所差异.滩脊的柽柳72.6%～95.4%水分来源于浅层地下水和含水量相对较高的深层土壤水(40～100 cm);高潮线附近的柽柳有40.7%～97.3%的水分来源于上层土壤水(0～40 cm),以避免海水和浅层地下水的盐分胁迫.柽柳对外界水盐条件变化具有较强的适应性,在海岸带可利用水资源缺乏的恶劣生境中具有更强的种间竞争优势,从而导致柽柳单优灌木群落的形成.     

山西省南方红豆杉自然分布与群落生态学特征

南方红豆杉为国家Ⅰ级重点保护植物,山西省东南部为其自然分布最北界.本文调查了南方红豆杉在山西省的野生分布,对其进行了群落划分、物种多样性、结构特征和竞争特性的研究.结果表明: 山西省南方红豆杉主要形成南方红豆杉+鹅耳枥群落、南方红豆杉-荆条群落、南方红豆杉-海州常山群落和南方红豆杉+栓皮栎群落,集中分布于陵川县磨河和阳城县蟒河自然保护区,其他地区均为散生;群落垂直结构明显,南方红豆杉已进入主林层,混交林林冠层高8~10 m,纯林高5～6 m;群落间的总体物种多样性指数和均匀度指数存在显著差异.南方红豆杉植株矮小,平均高5.16 m,乔木层、演替层分别占43.4%和56.6%,更新层缺乏;胸径＜16 cm的个体占67.6%,胸径32～40 cm的个体占4.0%,幼苗稀少,仅发现4株幼苗;大量的小径级个体虽可保证南方红豆杉在一定时间内维持稳定的种群结构,但幼苗缺失必将引起未来种群的衰退.山西省野生南方红豆杉分布相对集中,多数是小径级个体,导致其种内竞争激烈,种内竞争强度占61.8%;鹅耳枥和栓皮栎是研究地区的主要组成树种,对南方红豆杉造成的种间竞争压力最大.     

Plant regeneration from cell suspension-derived protoplasts of Primula malacoides and Primula obconica

Protoplasts were isolated from cell suspension cultures of Primula malacoides cv. ‘Lovely Tokyo’ and P. obconica cv. ‘Aalsmeer Giant White’. P. obconica protoplasts were embedded in 0.1% (w/v) gellan gum-solidified discs comprising MS medium supplemented with 3 mg/l of 2,4-D or picloram, 0.1 mg/l of zeatin, 0.2 M glucose and 0.2 M mannitol, and surrounded by a liquid medium of the same composition except for the addition of 0.1% (w/v) activated charcoal. The protoplasts formed visible colonies, which were transferred to the regeneration medium containing 30 g/l of sucrose, 0.1 mg/l of picloram and 2 mg/l of zeatin for shoot induction. P. malacoides protoplasts formed visible colonies when cultured in disc culture using 0.1% (w/v) gellan gum-solidified MS medium containing 5 mg/l of 2,4-D, 1 mg/l of NAA, 0.1 mg/l of zeatin and 0.4 M glucose. Small calli were transferred to MS medium supplemented with 5 mg/l of zeatin for shoot regeneration. The shoots of both species readily rooted on plant growth regulator-free 1/2 MS medium and successfully acclimatized to greenhouse conditions. The protoplast-derived plants showed some alterations in morphological characteristics from those of the in-vitro-germinated control plants.     

Fertile plant regeneration from protoplasts of a seed-propagated cultivar of Lilium × formolongi by utilizing meristematic nodular cell clumps

Plant regeneration from protoplasts of Lilium × formolongi cv. Azusa was achieved by utilizing suspension cultures of meristematic nodular cell clumps with a high plant regeneration ability. Creamy-white calli with embryogenic potential were initially induced from the seeds on Murashige and Skoog (MS) medium containing 4.1 μM picloram. The calli were then transferred into a liquid medium of the same composition, in which they turned into compact cell clumps which consisted of meristematic nodules. Protoplasts were readily isolated from these meristematic nodular cell clumps. Colonies were successfully formed from the protoplasts by embedding in 0.1% gellan gum-solidified MS medium containing 4.1 μM picloram and 0.5 M glucose. They regenerated shoots and roots on MS medium containing 2.2 μM benzylaminopurine (BAP). The plants thus obtained produced flowers with normal fertile pollen 8 months after successful transfer into soil. These plants had normal chromosome numbers (2n = 24) but had shorter leaves than original plants. They set seeds after as well as cross pollination.     

Accumulation of osmolytes and osmotic adjustment in water-stressed wheat (Triticum aestivum) and maize (Zea mays) as affected by calcium and its antagonists

Maize (Zea mays) and wheat (Triticum aestivum) were water stressed for 4 days at early vegetative growth (15-day-old) using PEG-6000 (−1.0 MPa), in the presence of 1 mM CaSO₄, 50 μM Verapamil (VP; calcium channel blocker); 50 μM Trifluoperazine (TFP; calmodulin antagonist) and then put to recovery in order to investigate the changes in osmoregulation in plants having C₃ and C₄ metabolism. Accumulation of proline (Pro) and quaternary ammonium compounds (QAC's), activities of pyrroline-5-carboxylate reductase (P5CR), proline dehydrogenase (PDH), water potential (Ψ_w), osmotic adjustment (OA), relative elongation rate (RER) and electrolyte leakage (EL) were examined during stress and recovery. Maize had significantly higher accumulation of Pro while wheat showed relatively more accumulation of QAC's. The activities of P5CR and PO were also significantly higher in maize than wheat. Maize shoots under stress showed higher Ψ_w, OA, RER and less EL than wheat shoots. Upon recovery from stress, maize regained its growth and water potential faster than wheat. Ca²⁺ elevated the accumulation of osmolytes in both the plants but OA was less sensitive to it. In the presence of Ca²⁺, wheat showed significantly more accumulation of osmolytes, higher Ψ_w, RER than maize. Ca²⁺ inhibitors partially reversed the effects of calcium indicating its involvement in governing solute accumulation. The differential sensitivity of maize and wheat towards water stress may be related to variation in endogenous calcium expression and its function.     

Asymmetric somatic hybridization between wheat (Triticum aestivum) and Avena sativa L.

Protoplasts from cell suspensions of young-embryo-derived calli, which were nonregenerable for long-term subculture and protoplasts from embryogenic calli with the regeneration capacity of 75% of the same wheat Jinan 177, were mixed as recipient. Protoplasts from embryogenic calli of Avena sativa (with the regeneration capacity of less than 10%) irradiated with UV at an intensity of 300 μW/cm² for 30 s, 1 min, 2 min, 3 min, 5 min were used as the donor. Protoplasts of the recipient and the donor were fused by PEG method. Many calli and normal green plants were regenerated at high frequency, and were verified as somatic hybrids by chromosome counting, isozyme, 5S rDNA spacer sequence analysis and GISH (genomic in situ hybridization). Fusion combination between protoplasts either from the cell suspensions or from the calli and UV-treated Avena sativa protoplasts could not regenerate green plants.     

Asymmetric somatic hybridization between Triticum aestivum L. and Haynaldia villosa Schur

Suspension cell-derived protoplasts of wheat, inactivated with different concentrations (0-2.5mol/L) of IOA, were fused by PEG method with the Haynaldia villosa protoplasts which originated from the calli 4-5d after subculture and were irradiated with 60Co-γ ray. Cell colonies, calli or regenerated plants were obtained from different combinations of fusion. The calli and plants were verified to be hybrids by chromosome counting, isozyme analysis and morphological inspection.     

不同施氮水平对冬小麦生长期土壤呼吸的影响

在目前全球氮沉降不断增加的背景下, 研究农田土壤呼吸对氮沉降的响应有助于理解未来生态系统碳循环对全球变暖的潜在影响。为探讨不同施氮浓度对华东地区冬小麦(Triticum aestivum)生长期土壤呼吸的影响, 该实验设计了对照组(不施加氮肥)和3种浓度施氮处理组(低浓度施氮15 g·m^-2·a^-1, 中等浓度施氮30 g·m^-2·a^-1, 高浓度施氮45 g·m^-2·a^-1)。使用便携式土壤CO₂通量观测仪LI-8100测定不同施氮浓度处理下冬小麦生长期(2013年12月至2014年5月)的土壤呼吸速率, 并探讨土壤呼吸与土壤温度、湿度等环境因素的关系。结果表明: 低、中、高3种浓度施氮处理的土壤呼吸速率平均值分别为5.29、6.17和6.75 μmol·m^-2 ·s^-1, 与对照组(土壤呼吸速率平均值为4.90 μmol·m^-2·s^-1)相比, 分别增加了7.8%、23.6%和37.8%; 地上生物量分别增加39.9%、104.4%和200.2%, 并与冬小麦生长季的总土壤呼吸正相关。5 cm深度土壤的温度与土壤呼吸速率呈指数关系(p < 0.05), 土壤呼吸季节变化的65%-75%由土壤温度引起, 其温度敏感性为2.09-2.32。结果表明, 添加氮肥促进了植物的生长, 增加了生物量, 从而增加了冬小麦农田的土壤呼吸速率。     

Intermediate fertile Triticum aestivum （＋）Agropyron elongatum somatic hybrids are generated by low doses of UV irradiation

We report the production and characterization of somatic hybrids between Triticum aestivum L. and Agropyron elongatum (Host) Nevishi (the synonym is Thinopyrum ponticum). Asymmetric protoplast fusion was performed between Agropyron elongatum protoplasts irradiated with a low UV dose and protoplasts of wheat taken from nonregenerable suspension cultures. More than 40 green plantlets were obtained from 15 regenerated clones and one of them produced seeds. The phenotypes of the hybrid plants and seeds were intermediate between wheat and Agropyron elongatum. All of the regenerated calli and plants were verified as intergeneric hybrids on the basis of morphological observation and analysis of isozyme, cytological, 5SrDNA spacer sequences and random amplified polymorphic DNA (RAPD). RFLP analysis of the mitochondrial genome revealed evidence of random segregation and recombination of mtDNA.     

Asymmetric somatic hybridization between wheat (Triticum aestivum) and Avena sativa L.

Common wheat is one of the most important cereal crops in the world. The improvement of its yield and quality by the introduction of heterologous gene(s) is very significant. Avena sativa L. (2n = 42), belonging to the Avena tribe, possesses resistance to drought, coldness and many dis-eases. Its contents of proteins and fat in seed, especially lysine and unsaturated fatty acid are highest in crops, therefore it is regarded as healthy food. Sexual hybridization between wheat and Avena sativa…     

Asymmetric somatic hybridization between wheat (<Emphasis Type="Italic">Triticum aestivum</Emphasis>) and <Emphasis Type="Italic">Avena sativa</Emphasis> L.

Protoplasts from cell suspensions of young-embryo-derived calli, whichwere non- regenerable for long-term subculture and protoplasts from embryogenic calli with the regeneration capacity of 75% of the same wheat Jinan 177, were mixed as recipient. Protoplasts from embryogenic calli of Avena sativa (with the regeneration capacity of less than 10%) irradiated with UV at an intensity of300 μW/cm2 for 30 s, 1 min, 2 min, 3 min, 5 min were used as the donor. Protoplasts of the recipient and the donor were fused by PEG method. Many calli and normal green plants were regenerated at high frequency, and were verified as somatic hybrids by chromosome counting, isozyme, 5S rDNA spacer sequence analysis and GISH (genomic in situ hybridization). Fusion combination between protoplasts either from the cell suspensions or from the calli and UV-treated Avena sativa protoplasts could not regenerate green plants.     

水平结构配置对冬小麦冠层垂直结构、微环境及产量的影响

作物的冠层结构是影响产量的重要因素,群体微环境反映了作物冠层内小气候的变化,与作物的冠层结构及产量形成密切相关。该研究在大田试验条件下,设置等行距(R₁,20 cm+20 cm)、宽窄行(R₂,12 cm+12 cm+12 cm+24 cm)两种不同行距和低(D₁,120.0 kg·hm^–2)、中(D₂,157.5 kg·hm^–2)、高(D₃,195.0 kg·hm^–2) 3个播量配置组合,分析了不同处理组合下冬小麦(Triticum aestivum)生育后期冠层垂直结构、群体微环境及产量表现,旨在优化小麦绿色栽培措施,在不增加水肥投入情况下,挖掘冬小麦的生产潜力和进一步增产的可能性和可行性。结果表明:冬小麦上、中、下3个层次冠层开度(DIFN)、平均叶倾角(MLA)及叶面积指数(LAI)均表现为R₂大于R₁,且R₂行距上层和中层DIFN、各层次...     

Somatic embryogenesis and plant regeneration from protoplasts isolated from embryogenic cell suspensions of wheat (Triticum aestivum L.)

We describe the early formation of somatic embryos followed by plant regeneration from protoplasts isolated from an embryogenic wheat cell suspension, which was initiated from small granular (0.2 to 1 mm in size) embryogenic calli. These granular calli formed embryogenic cell suspensions within 20 days in liquid culture, and were selected gradually from young inflorescence-derived nodular embryogenic calli of the winter wheat cv. Kehong 1041. The division frequency of protoplasts was 11 to 16%, and the frequency of differentiation into plants was about 0.001% (number of plants formed divided by the total number of protoplasts plated). About 20% of somatic embryos present in the culture formed directly from protoplast-derived cells within 15 days of cultures.     

外源水杨酸对高温胁迫下金莲花幼苗生理及电阻抗参数的影响

为解决金莲花幼苗在北方低海拔地区越夏过程中生长不良的问题，研究喷施水杨酸对高温胁迫下金莲花幼苗生理及电阻抗参数的影响，为提高金莲花的耐热性提供理论依据。以金莲花幼苗为试验材料，外源水杨酸（SA）设置4个浓度梯度（0.5、1.0、1.5、2.0 mmol·L^-1），喷施处理后置于人工气候箱内进行昼38℃/夜30℃的高温胁迫试验。结果表明：喷施0.5~1.0 mmol·L^-1水杨酸能有效降低金莲花幼苗的热害指数、相对电导率和丙二醛（MDA）含量，缓解叶绿素降解、显著提高可溶性糖含量、游离脯氨酸含量以及超氧化物歧化酶（SOD）活性，其中以1.0 mmol·L^-1的水杨酸（SA）处理效果最好，说明喷施适宜浓度的SA可以提高金莲花的耐热性，从而缓解高温伤害。相关性分析表明，热害指数与相对电导率呈极显著正相关（P<0.01），与叶绿素总量、可溶性糖含量胞外电阻率（r_e）呈显著负相关（P<0.05），胞内电阻率（r_i）与Pro含量呈显著正相关，弛豫时间（τ）与Pro含量和SOD活性呈极显著正相关，因此胞外电阻率、胞内电阻率和弛豫时间等电阻抗参数可作为金莲花对高温逆境反应的评价指标。     

小麦根系特征对干旱胁迫的响应

干旱胁迫时, 小麦(Triticum aestivum)根系率先产生应激响应, 同时向地上部发出信号, 诱导地上部发生生理反应, 从而提高植株抗旱能力。根系构型包括平面几何性状和立体几何结构(即拓扑构型), 具有遗传稳定性和可塑性。干旱胁迫影响根系理化特性, 如根源化学信号、根系细胞酶类和根系渗透作用的响应。根系通过调整其解剖学结构和水分吸收动力等来适应干旱胁迫。该文从根系构型、理化特性和解剖学结构3个方面, 系统阐述了小麦根系特征对干旱胁迫的响应, 并探讨了其与干旱胁迫的关系和当前研究中存在的问题, 以期为相关研究提供参考。     

Stably transformed callus of wheat by electroporation-induced direct gene transfer

Fertile plants of wheat have been regenerated from protoplasts in several laboratories. The objective of this study was to develop a transformation system using protoplasts as target cells. Protoplasts were isolated from cell suspensions initiated from an anther-derived callus. The protoplasts were transformed by electroporation using pBARGUS or pBAS, both carrying the Basta resistance (BAR) gene. A total of 2,761 calli were produced from electroporation transformed protoplasts in 3 independent experiments. Six calli survived selective culture on 10 mg/l phosphinothricin (PPT), a concentration that completely inhibited the growth of non-transformed wheat callus. Five PPT resistant calli showed phosphinothricin acetyltransferase (PAT) activity, whereas the sixth probably was a mutant. The transformed wheat calli could tolerate PPT concentrations up to 2,560 mg/l. Southern blot analyses confirmed the integration of the BAR gene in wheat genomes. The integrated DNA sequence may have partially methylated and tandemly repeated at least once. These results demonstrate the production of stably transformed wheat calli by electroporation-mediated direct gene transfer into protoplasts.     

小麦根系特征对干旱胁迫的响应

干旱胁迫时, 小麦(Triticum aestivum)根系率先产生应激响应, 同时向地上部发出信号, 诱导地上部发生生理反应, 从而提高植株抗旱能力。根系构型包括平面几何性状和立体几何结构(即拓扑构型), 具有遗传稳定性和可塑性。干旱胁迫影响根系理化特性, 如根源化学信号、根系细胞酶类和根系渗透作用的响应。根系通过调整其解剖学结构和水分吸收动力等来适应干旱胁迫。该文从根系构型、理化特性和解剖学结构3个方面, 系统阐述了小麦根系特征对干旱胁迫的响应, 并探讨了其与干旱胁迫的关系和当前研究中存在的问题, 以期为相关研究提供参考。     

Somatic hybrids of Datura innoxia Mill.+Datura discolor Bernh. and of Datura innoxia Mill.+Datura stramonium L. var tatula L.

Summary Following fusion of protoplasts from a chlorophyll-deficient diploid mutant of Datura innoxia Mill. which can be regenerated to shoots, with green wild-type protoplasts of Datura stramonium L. var. tatula L. which can not, it was possible to isolate 49 green hybrid calli on agar medium. Most of these somatic hybrid calli gave rise to leaves and shoots. The chromosome numbers of the somatic hybrids were determined: 15 were tetraploid (amphidiploid), 24 hexaploid, and the other showed an aneuploid chromosome number.In a similar experiment protoplasts of the Datura innoxia mutant were fused with green wild-type protoplasts of Datura discolor Bernh. which are also not able to be regenerated, four green calli were obtained from which leaves and shoots developed after some transfers on agar medium. Three of them showed the amphidiploid (48) chromosome number, whereas one possessed an aneuploid number of 46 chromosomes.After transfer of rooted shoots to soil flowering plants could be obtained in both combinations. The habits of the somatic hybrids in both combinations were intermediate between the habits of the respective parental plants.Dedicated to my father, Prof. Dr. Theodor Schieder, on the occasion of his 70th birthday.     

黄河三角洲盐碱地不同植被模式的土壤改良效应

黄河三角洲是我国滨海盐碱地的重要分布区,种植植被是盐碱地绿色改良的主要生态修复措施。为探讨滨海盐碱地不同植被模式的土壤改良效应,探索适宜植被模式,选取黄河三角洲盐碱地竹柳+NyPa草、旱柳+NyPa草、柽柳+紫花苜蓿、白蜡+柽柳+紫花苜蓿4种林草措施为研究对象,以纯竹柳为对照,测定土壤水分物理参数、盐碱含量、土壤养分及微生物数量等20个指标,并利用主成分分析、聚类分析和模糊数学隶属函数等统计方法评价了不同植被模式的土壤改良效应。结果表明: 林草复合模式可显著改善滨海盐碱地的土壤理化性能,增加土壤孔隙度和贮水量,降低土壤密度,提高土壤有机质、速效养分含量和土壤微生物数量。其中,白蜡+柽柳+紫花苜蓿的乔灌草混交模式在压碱抑盐、增加土壤养分和微生物数量的效果最好,而旱柳+NyPa草的乔草混交模式改良土壤水分物理性能的效果最好。不同植被模式对黄河三角洲滨海盐碱地的综合改良效应表现为白蜡+柽柳+紫花苜蓿>旱柳+NyPa草>竹柳+NyPa草>柽柳+紫花苜蓿。