Variation in seed morphometric characters,oil content and azadirachtin content of seeds,in vitro shoot cultures and callus cultures among different populations of <Emphasis Type="Italic">Azadirachta indica</Emphasis>

We studied seed morphometric characters, oil content and azadirachtin content in a medicinally important and insect repellent tree species Azadirachta indica A. Juss. Ten populations were selected from two central Indian states of Madhya Pradesh and Chhattisgarh and twenty candidate plus trees were selected from each population based on tree attributes. Azadirachtin content was estimated in seeds, in vitro shoots and callus cultures through HPLC. Significant variation (p < 0.05) was observed in seed morphometric characters, oil content and azadirachtin content among the ten populations. Maximum value for 100 seed weight was obtained in seeds of Raigarh (30.51 g) which was on par with seeds of Bargi (28.37 g) and Chhatarpur (26.36 g). The ratio of highest kernel to seed was also obtained in seeds of Raigarh (60.11%). Kernels from Chhatarpur population recorded the highest oil content (24.20%), which was equal to that in Sihore (23.75%) and Katni (21.75%). We recorded highly significant variation in azadirachtin content in seed samples, in vitro shoot samples and callus cultures. Maximum azadirachtin was present in the seeds of Gwalior (0.882%) which was on par with azadirachtin content in seeds of Chhatarpur, Katni, Sihore, Shahdol and Chhindwara. Maximum azadirachtin in the in vitro shoots was present in the cultures of Gwalior (0.218%) which was statistically on par with Sihore (0.176%). Similarly, in callus cultures of nodal segments maximum azadirachtin was recorded in the Gwalior population (0.033%) which was statistically on par with Sihore (0.028%). The trees of Gwalior and Sihore populations can be produced in large numbers through vegetative propagation to obtain seeds with higher azadirachtin content.     

Screening <Emphasis Type="Italic">Azadirachta indica</Emphasis> tree for enhancing azadirachtin and oil contents in dry areas of Gujarat,India

Azadirachta indica seed samples were collected from four different agro-ecological regions(AERs) viz.,AER-2,AER-4,AER-5A and AER-5B of Gujarat state,India during 2000 to 2002 with an aim to assess variability in azadirachtin,oil and fatty acids content of the seeds and assess correlation of these parameters with morphological characters.Azadirachtin,oil and fatty acids content indicated significant(p<0.01) variations between years and AERs in Gujarat.The azadirachtin ranged from 142 to 9 527 μg·g-1 seed kernel with an average of 2 426 μg·g-1 for the state.AER-5B recorded highest azadirachtin and oil content.Fatty acid composition was found to be affected by environmental factors particularly varying degree of annual rainfall and temperature during fruit ripening period.Conclusively trees growing in AER-5B performed better in term of azadirachtin,oil and stearic acid content.Tree girth at breast height showed no significant relation with these biochemicals.     

印楝素含量与种子形态及成熟期关系的初步研究

对印楝素含量与种子形态表征及成熟期的关系进行了初步研究.种子的长宽比与印楝素含量相关,并且认为它是能反映印楝素含量的主要表征,得出种子长宽比(X)与印楝素含量(Y)的关系为:Y=0.295 0.134X-0.028X2.种子长宽比接近2.39时,种仁的印楝素含量出现峰值,可将印楝种子的形态表征作为初步估测印楝素含量的指示参数,而种子的成熟期与印楝素含量无关.在印楝原料林良种优良母树筛选中应尽量选择各个种源中生长良好、结实量大、种子饱满且长宽比接近2.39的植株.     

Provenance variations in seed-related characters and oil content of <Emphasis Type="Italic">Calophyllum inophyllum</Emphasis> L. in northern Australia and Sri Lanka

Seed morphometric characters and oil content were studied in multiple-use plant, Calophyllum inophyllum L. of two countries, Australia (southern hemisphere) and Sri Lanka (northern hemisphere). Seven provenances were selected which included three from northern Australia and four from Sri Lanka. Twelve Candidate plus trees (CPTs) each were selected from 2 to 3 different locations within each provenance based on the morphometric and qualitative traits (GBH > 100 cm). Seed collection in both hemispheres was carried out from May to August 2008. Calophyllum inophyllum L. provenances showed a distinct hemisphere variation in their seed-related characters and oil content. All provenances differed significantly (P < 0.05) with one another in seed length. Seeds from Anuradhapura, Sri Lanka and seeds from Cardwell Australia recorded the highest (≈57%) and the lowest oil (≈31%) content respectively. Strong correlations were found between seed morphometric characters. Relatively weaker correlations were found between seed morphometric characters and oil content. Variations in seed-related characters were largely attributed by the provenance contribution which shows the significance of the effect of genetic variability on above mentioned seed related characters.     

Variation in Seed and Seedling Traits of Celtis australis, a Multipurpose Tree, in Central Himalaya, India

Seeds of Celtis australis were collected from 13 different sources, ranging from 550 to 1980 masl, in Central Himalaya, India. Significant (p = 0.05) variations were observed for seed traits among provenances. However, among various characters, seed weight exhibited maximum variation between seed populations compared to other morphological characters. Between provenances, seed weight ranged from 47.8 to 83.1 g/1000 seed, with mean value of 66.9 ± 10.7 g/1000 seed. Significant (p = 0.01) positive correlation was found between morphological characters of seeds including seed weight and elevational range of seed source. For one year old seedlings, average shoot and root growth was 61.1 ± 13.3 and 30.5 ± 5.4 cm, respectively, irrespective of provenance variation. Inter-comparing biomass yield of the seedlings with altitude, average biomass production was 8.4 ± 2.5, 9.4 ± 3.3 and 12.7 ± 1.7 g/plant, respectively, for low (550–1000 masl), middle (1050–1250 masl) and high (1350–1980 masl) altitudinal populations. Significant (p = 0.01) positive correlation between growth performance of seedlings and altitude of the seed source was recorded. Across the provenances, shoots had the highest proportion of total biomass (42.3%), followed by leaves (32.6%) and roots (24.6%). Among various provenances, Badiyargaon, Agroda, Guptakashi, Jakholi, Gajeli, Srinagar and Palampur populations produced heavier seedlings and grew faster compared to seedlings of other sources.     

西部地区文冠果种群种子特征及主要化学成分的地理变化

通过野外实际调查和测定,对我国西部地区8个文冠果天然群体的种子特征及主要化学成分进行研究。结果表明:8个文冠果种群的种子横径为1.05～1.51cm,纵径为0.86～1.38cm,种形指数为1.07～1.32,种群间存在显著差异;陕西延安种群种子的千粒质量和出仁率均最大,分别为1108.4g和48.28%,显著高于甘肃和青海种群;8个种群的种仁含油率均在53%以上,且不同种群间差异显著;文冠果种仁油含有12种脂肪酸,其中含量最高的是亚油酸(40.90%～46.12%),其次是油酸(29.21%～31.25%);含有17种氨基酸(22.53%～27.58%),包括7种必需氨基酸,占总氨基酸的27.61%。相关分析结果表明:文冠果种形指数表现出明显的地理变化规律,随经度的增加,种形指数逐渐减小;种子千粒质量与4—9月份的降水量呈极显著正相关,与年平均温度呈显著正相关,与年干燥度呈显著负相关;种仁油含量随地理经度的减小而降低,与4—9月份的降水量呈显著正相关;种仁氨基酸含量与地理经度及无霜期相关性显著,随经度和无霜期的增加而增大,即东部地区氨基酸含量较高。     

Rapid in vitro cloning of a 40-year-old tree of <Emphasis Type="Italic">Azadirachta indica</Emphasis> A. Juss. (Neem) employing nodal stem segments

An efficient in vitro process for rapid clonal propagation of a 40-year-old tree of Azadirachta indica employing nodal stem segments was developed. Season of collection and maturity of explants showed direct influence on bud-break. Nodal stem segments collected during the month of April gave best response. Maximum bud-break (78.6–81%) was obtained when middle order nodes (3rd or 4th node from apex) were taken. Amongst different cytokinins used, 6-benzylaminopurine (BAP) at the concentration of 1.11 μM was found most effective in inducing multiple shoots, whereas inorganic and organic constituents of the medium influenced growth and general condition of proliferating shoots. On an average 3.1 shoots per explant were regenerated in modified Murashige and Skoog medium supplemented with 1.11 μM BAP, 1.43 μM indole-3-acetic acid (IAA) and 81.43 μM adenine hemisulphate. Isolated shoots were rooted in presence of 2.46 μM indole-3-butryic acid (IBA). Root induction took place in 8–10 days with 100% rooting. The in vitro-raised plantlets were successfully transplanted in potted soil and finally grown under field conditions with 100% survival. The genetic fidelity of such in vitro-raised field-grown plants was ascertained by random amplified polymorphic DNA (RAPD) markers. Furthermore, the azadirachtin content of in vitro-cloned plants was found comparable to the mother tree proving their chemical stability also. The protocol developed holds good for in vitro cloning of mature elite neem trees.     

几种因素对东革阿里的愈伤组织中抗肿瘤药物成分累积的影响(英文)

研究提高东葛阿里的愈伤组织中抗肿瘤药物成分的累积。有些因素(不同培养基成分和物理因素)影响东葛阿里的愈伤组织中抗肿瘤药物成分的积累。结果表明:在1/4MS基本培养基的愈伤组织中获得较高的抗肿瘤药物成分(3.84mg·g-1干重)。与在基本培养基中添加碳源相比,添加2%果糖,抗肿瘤药物成分产量最高(4.59mg·g-1)。添加激素(3,6-Dichloro-o-aniscicacid,2.0mg·L-1),也可提高抗肿瘤药物成分产量(12.3mg·g-1)。在基本培养基pH值为5.5时,可获得较高的抗肿瘤药物成分产量(1.53mg·g-1)。当基本培养基附加苯丙氨酸(1×10-1μM)时,抗肿瘤药物成分产量也被提高到2.34mg·g-1。成功地提取了在试管培养下获得的抗肿瘤药物成分,具有大范围生产的潜力。     

Target seed moisture content,chilling and priming pretreatments influence germination temperature response in Alnus glutinosa and Betula pubescens

Abstract

The response of common alder [Alnus glutinosa (L.) Gaertn.] and downy birch (Betula pubescens Ehrh.) seeds to germination temperature was examined following chilling and priming. Seeds of two seed lots of each species were subjected to combinations of chilling (4±1°C) and priming (20°C) treatments in fully imbibed (FI) state or a lower target seed moisture content (TMC) level (30% and 35% in alder and birch, respectively). After treatment, the seeds were allowed to germinate for 56 days at constant temperatures of 7.5, 10, 15, 20, 25 or 30°C. The response to temperature and pretreatment differed between species, but the effect was consistent in each seed lot within each species. In alder, the TMC seeds germinated well across the full range of temperatures, whereas there was an optimum temperature (22–23°C) for seeds given the FI pretreatment. Priming had no significant effect on the germination response of the TMC seeds in alder, but priming greatly improved germination in the FI seeds, especially at the lower germination temperatures (optimum 18–19°C). In contrast, in birch, the TMC seeds germinated better across the full temperature range, but the optimal germination temperature (15°C) was the same for all seed pretreatments. Priming improved germination in both the FI and TMC seeds in birch.     

Biochemical investigations during in vitro adventitious shoot regeneration in leaflet explants from nodal segments of a mature Albizia procera tree

The in vitro adventitious shoot differentiation in leaflet explants of an adult tree differed from that of leaflet explants of seedlings of Albizia procera(Roxb.)Benth. reported previously elsewhere. The leaflet explants from an adult tree passed through an initial callus phase for30 days on MS medium supplemented with 3 % sucrose,2.5 l M 2,4-D followed by a subsequent adventitious shoot differentiation phase for another 30 days on half MS medium supplemented with 0.25 l M each of BA and IBA.The regeneration rate of in vitro adventitious shoots in explants from the adult tree, i.e.1.66 shoots/callus, was lower than that from seedlings, i.e. [10 shoots/callus,which was reported elsewhere. Correspondingly, the activities of nitrate reductase and peroxidase, and endogenous phenol content remained very low during in vitro adventitious shoot differentiation in leaflet explants of an adult tree possibly due to lower availability of competent stem(juvenile) cells for the process.     

芍药种子含油率与脂肪酸成分研究

[目的]本文对芍药种子含油率、脂肪酸成分及含量进行了探讨,为芍药开发成油料植物提供参考。[方法]选取芍药36个栽培品种和1个野生居群为样本,采用索氏提取法提取种子油脂并测定含油率,用气相色谱—质谱联用仪(GC-MS)对提取的籽油成分进行分析。[结果]表明:芍药36个栽培品种和1个野生居群种子平均含油率为20.20%。芍药籽油主要含有豆蔻酸、棕榈酸、棕榈一烯酸、硬脂酸、油酸、亚油酸、亚麻酸和花生酸8种脂肪酸,其中,亚麻酸含量最高,平均达34.14%,其次为油酸和亚油酸,这3种不饱和脂肪酸的含量占全部脂肪酸的93.37%。芍药籽油脂肪酸含量变化相对稳定,各脂肪酸成分的变异系数从大到小依次为亚油酸(13.90%)油酸(13.51%)亚麻酸(10.71%)棕榈酸(0.18%)硬脂酸(0.09%)花生酸(0.002%)、棕榈一烯酸(0.002%)豆蔻酸(0.00%)。[结论]芍药种子含油率高,富含亚麻酸、亚油酸和油酸,且含量相对稳定,变异很小,具有开发为高含量亚麻酸食用油潜力。     

Effect of organogenesis on steroidal saponin biosynthesis in calli cultures of Ruscus aculeatus

Calli cultures derived from Ruscus aculeatus rhizomes were investigated for their potential to biosynthesize saponins. The capacity of undifferentiated tissues to form steroidal saponins is very limited, but when the calli developed organogenesis, mainly aerial shoots and roots, the saponin production increased significantly. Plantlets regenerated from aerial shoots of Ruscus calli showed a saponin pattern similar to that of the callus cultures but the levels of saponins found in the aerial part and roots were significantly greater.     

Mevalonate kinase activity during different stages of plant regeneration from nodular callus cultures in white pine (Pinus strobus)

Mevalonate kinase (MK) catalyzes a step in the isoprenoid biosynthetic pathway, which leads to a huge number of compounds that play important roles in plant growth and development. Here, we report on changes in MK activity in white pine (Pinus strobus L.) during plant regeneration by adventitious shoot organogenesis from cotyledons of mature embryos, including nodular callus induction, shoot formation and rooting. Nodular calli were induced from Pinus strobus (PS) embryos by culture in nodular callus induction medium in a 0-, 8- or 16-h photoperiod. Mevalonate kinase activity peaked in nodular calli after three weeks of culture on nodular callus induction medium in a 16-h photoperiod, whereas frequency of nodular callus formation peaked after 4 weeks of culture on nodular callus induction medium in darkness. During adventitious shoot formation, MK activity peaked in shoots derived from dark-grown nodular calli after 3 weeks on bud formation medium, and frequency of shoot formation was highest in dark-grown nodular calli cultured on bud formation medium for 4 weeks. During rooting, MK activity peaked 2 weeks after transfer of adventitious shoots to rooting medium and rooting frequency was highest in adventitious shoots after 3 weeks on rooting medium. Although during nodular callus induction in darkness MK activity was inversely related to frequency of nodular callus formation, MK activity was highly correlated with frequency of shoot formation and with rooting frequency. The observed increase in MK activity preceding rooting suggests that MK could serve as a marker for rooting of white pine shoots in vitro.     

Seed Development, Maturation and Storage Behaviour of Mimusops elengi L.

Mass maturity (end of the seed-filling phase) occurred at about 72 days after flowering (DAF) in developing seeds of Mimusops elengi, at which time seed moisture content had declined to about 55%. The onset of ability to germinate was detected at 56 DAF and seeds showed 98% germination by 84 DAF. Tolerance of desiccation to 10% moisture content was first detected at 70 DAF and was maximal by 84 DAF. Delaying collection by a further 14 days to 98 DAF, when fruits began to be shed, reduced seed viability, particularly for seeds first dried to 10% moisture content. Hence the best time for seed collection appears to be about 14 days before fruits shed. In a separate investigation with six different seed lots, desiccation below about 8–12% moisture content reduced viability (considerably in some lots). The viability of dry seeds (below about 10% moisture content) stored hermetically was reduced at cool temperatures (5 °C and below), and none survived storage at sub-zero temperatures. The results suggest that Mimusops elengi shows intermediate seed storage behaviour and that the optimal hermetic seed storage environment is about 10% moisture content at 10 °C, while short-term, moist, aerated storage at high (40%) moisture content is also feasible.     

Interactions between callus cultures of Pinus sylvestris and pine fungi with different trophic properties

Fungal virulence may be studied using tissues cultures of host plants in dual cultures in vitro, enabling analyses of interactions with undifferentiated cells of their host plants. Three genotypes of Pinus sylvestris callus, initiated by somatic embryogenesis, were used for establishing dual cultures with fungi pathogenic, endophytic or saprotrophic on pine needles or shoots. Fungal growth towards the plant callus tissue differed, depending on the life strategy of the fungus. The pathogen Gremmeniella abietina proved the slowest colonizer of callus whereas the saprotrophic Phacidium lacerum was the fastest. Gremmeniella abietina partially overgrew the callus, causing extensive necrosis and death within 10 days after inoculation. Anthostomella formosa, an endophyte of pines, did not cause evident symptoms of callus degradation: after 10 days of dual culture, the callus cells remained greenish and at least 50% of cells were alive. In dual cultures Ph. lacerum, callus remained alive until the end of the experiment, maintaining a white‐creamy colour with a loose cell structure. Electrophoresis of protein extracts from the callus showed the presence of additional bands of 25–35 kDa only in host tissues challenged with the pathogen G. abietina, possibly indicating the production of pathogenesis‐related proteins. This work has shown that pine callus does not respond equally to challenge with different fungal isolates. In general, one‐third of the isolates of each fungus examined showed greater virulence compared to other isolates.     

Induction of flavonoid production by UV-B radiation in Passiflora quadrangularis callus cultures

Callus cultures from several species of Passiflora were initiated in vitro, and their capacity to produce four glycosyl flavonoids (orientin, isoorientin, vitexin, isovitexin) was analysed. The aim of the present work was to examine the possible role of UV-B irradiation and elicitation with methyl jasmonate (MJ) on the production of these compounds in callus cultures. All the species tested (P. incarnata, P. quadrangularis, P. edulis) formed friable callus from leaf explants after 4 weeks on medium supplemented with kinetin and 2,4-dichlorophenoxyacetic acid. Among them, P. quadrangularis turned out to have a faster growth rate and a more friable texture, and was therefore chosen for experiments with elicitors. In callus cultures only small amounts of isoorientin were found, while the concentration of the other flavonoids was below the detection limit. UV-B irradiation of calluses was able to increase the production of all four glycosyl flavonoids. After a 7-day exposure of cultures to UV-B light, the production of isoorientin reached concentrations similar to those found in fresh leaves from glasshouse-grown plants. Elicitation with methyl jasmonate also enhanced orientin, vitexin and isovitexin concentrations, even though the stimulation was about 6-fold weaker for orientin and vitexin and about 40-fold for isovitexin, than that exerted by UV-B treatment. Callus cultures treated with the UV-B dose which most enhanced flavonoid production showed a higher antioxidant activity compared to untreated calluses, with an increase ranging from 28% to 76%. Results show that the secondary metabolite biosynthetic capacity of Passiflora tissue cultures can be enhanced by appropriate forms of elicitation.     

Germination requirements of <Emphasis Type="Italic">Allanblackia parviflora</Emphasis> seeds and early growth of seedlings

Allanblackia parviflora A. Chev, is an indigenous fruit tree species that could be used in agroforestry systems with both environmental and economic benefits. The seed oil is of prime importance as a foreign exchange earner and is being developed as a rural based enterprise in many African countries notably Ghana, Nigeria, Cameroon and Tanzania. Methods for propagation and conservation of the species are therefore of prime importance. The seeds of the species are however difficult to germinate hindering its domestication process. Allanblackia parviflora seeds were collected from seven different populations in Ghana and were subjected to four different seed germination trials at the nursery of CSIR-Forestry Research Institute of Ghana at Fumesua, Kumasi. This was followed by studying the effect of stage of seed germination on establishment after potting. Intact seeds began germination at 7 months after sowing and at 24 months, significant (P < 0.001) variations were observed in the seed germination ability of 74 accessions. Mean seed germination ranged from 0 to 35%. Significant differences in seed germination percentages among populations (P < 0.001) and provenances (P < 0.05) were also observed. Removal of seed coat significantly (P < 0.001) enhanced seed germination. Germination percentages of seeds with seed coat removed ranged from 6 to 43% while seed germination percentages for intact seeds were less or equal to 3%. Incubation of seeds with seed coat removed in polythene bags gave an added advantage. Seeds with seed coat removed and kept in polythene bags started germinating from 2 weeks and within 10 months, 75 and 68% germination were obtained for plane and black polythene bags, respectively. Removal of germinants and potting when shoot length was at least 1 cm gave significantly higher growth (P < 0.001 for shoot height, P < 0.05 for stem diameter, P < 0.001 for number of leaves produced) compared to when only radicle had emerged. It became evident in this study that seeds should be classified as fully germinated only when shoots begin to grow.     

Morphogenesis in leaf and single-cell cultures of mature Juniperus oxycedrus

Single cells were mechanically isolated from leaf-derived callus of mature Juniperus oxycedrus L. These cells divided and gave rise to callus when plated on medium containing growth regulators. Best plating efficiency was obtained on a modified Schenk and Hildebrandt medium supplemented with 0.6 micro M 2,4-dichlorophenoxyacetic acid and 100 mg l(-1) casein hydrolyzate. Although single-cell-derived callus showed poor morphogenic potential, both adventitious shoots and embryogenic tissues differentiated from the callus. We also achieved induction of somatic embryogenesis in leaf explants of mature J. oxycedrus trees cultured in the presence of 6.0 or 10.0 micro M 2,4-dichlorophenoxyacetic acid or picloram. Frequency of embryogenic callus ranged from 6 to 18%; however, under the culture conditions tested, isolated embryos failed to develop into plants.     

竹子的离体培养研究

近２０ａ来已对２０个属７０余种竹子进行离体培养研究,以侧芽,顶芽,成熟胚作外植体诱导愈伤组织,由愈伤组织制备悬浮细胞进行细胞悬浮培养,由悬浮细胞制备原生质体进行原生质体培养。竹子愈伤组织经不定芽途径或体细胞胚发生途径再生完整植株。通过芽尖培养增殖新生芽进行竹微繁殖,并获得脱病毒种苗。以芽为外植体增殖的新芽或组织再生苗经继代培养诱导竹试管开花结实。     

圆齿野鸦椿籽油提取工艺及脂肪酸组分分析

为高效利用圆齿野鸦椿籽油,采用正己烷加热回流提取圆齿野鸦椿籽油,分别考察了提取时间、提取温度和正己烷-圆齿野鸦椿籽粉末比(液料比)对圆齿野鸦椿籽油提取得率的影响.以单因素试验的结果为依据,运用Design-Expert 8.0.6软件进行Box-Behnken响应面设计及数据分析,进一步对回流提取圆齿野鸦椿籽油的工艺进...