刺楸种子层积过程中内源植物激素的动态变化及激素在种子休眠中的作用

本文主要以高效液相色谱为主要手段,结合生物测定方法,测定了4种不同层积条件下激素动态变化。结果表明刺楸干种子中存在有两种抑制物质-脱落酸(ABA)和香豆素(C),在种子层积的不同阶段又相继有GA₃,IAA和Z出现,并在层积后熟过程中呈现非常有规律的变化。根据激素的变化,可把种子整个后熟过程分三个阶段;即阶段Ⅰ,以抑制物质(ABA,C)和IAA水平迅速减少为主要特征,阶段Ⅱ主要表现为GA₃和Z合量的上升,阶段Ⅲ各种激素处于相对稳定的状态。种子的休眠与否可能主要取决于阶段Ⅱ的状况。在刺楸种子胚形态后熟期间,胚的生长与分化同ABA和C水平有很高的相关性,但同时也受GA₃和IAA的调节。生理后熟主要与Z有关,同ABA和C无明显相关性。同时本文还对激素相对水平做了初步研究,发现GA₃/ABA+IAA,Z/GA₃+IAA和GA₃/C+IAA^*,在种子后熟期间的变化同胚生长发育存在高度的一致性。认为激素的相对水平对种子休眠起重要的控制作用,还推测激素的作用可能类似于"板机"机制。     

野鸦椿种子内源抑制物活性初探

以野鸦椿(Euscaphis japonica)种壳和胚为材料,甲醇浸提得到种壳浸提液和胚浸提液,配制浸提液浓度梯度为原浸提液浓度的10%、20%、30%、40%,研究不同浓度的种壳和胚浸提液对白菜、小麦、绿豆种子底物酶活性、发芽率、幼苗根长和苗高的影响,萃取和分离种壳和胚甲醇浸提液中的内源抑制物质,探讨野鸦椿种子内源抑制物质的活性与成分。结果表明:随着种壳和胚浸提液浓度的增加,白菜种子酸性磷酸酶活性和发芽率均显著降低(P0.05),表现为抑制作用递增,种壳的抑制作用小于胚,而幼苗的根长和苗高则表现为低促高抑,在10%浸提液处理下根长和苗高达最大值,种壳的促进效果弱于胚;小麦种子淀粉酶活性及幼苗的根长和苗高递减(P0.05),表现为抑制作用递增,而发芽率则在浓度≤20%时差异不明显(P0.05),浓度为30%时显著降低(P0.05),40%时发芽率为0,种壳的抑制作用大于胚;绿豆种子蛋白酶活性、发芽率、幼苗根长和苗高均在浸提液浓度≥20%时显著下降(P0.05),且种壳的作用效果小于胚。种子内源抑制物萃取及分离表明,外壳中含酚酸类较胚多,含碱类较胚少。综上认为,野鸦椿种壳和胚中均含有较高活性的内源抑制物,但性质、成分及含量存在差异,外壳内源抑制物主要作用对象为淀粉类物质,胚乳内源抑制物主要作用于油脂类和蛋白类物质。     

种子发芽的抑制物质

许多植物的种子都有休眠现象,引起休眠的原因不一。在排除了胚未成熟,种皮机械障碍等因素后,如果给以适当的温、光、气、水条件仍不萌发,则应考虑是否有发芽抑制物的存在。发芽抑制物的种类抑制种子发芽的物质很多,既有简单的无机物,也有许多有机物及植物激素。氢氰酸（HCN）是常见的发芽抑制物,主要以耷的形式分布于植物界中。如苦费科中的梅、杏等,其种子中的苦杏仁耷含量可达百分之几。银杏、甘薯、高粱、亚麻等种子中均含有抑制种子萌发的各种氨氰酸的着类。如果在播有小麦种子的培养血中放两个苦杏仁,小麦种子发芽就受到抑…     

开花前后子房中生长调节物质的变化及其与结果及单性结果的关系

本研究由桔子、柿子(包括有核品种与单性结果品种)及南瓜(有籽品种)在开花前后的子房中提取出酸性乙醚溶解物质,并用小麦芽鞘垂直生长法测定其活性。结果表明。在所测定的各品种中开花前子房中的生长促进物与生长抑制物的活性都很高。但在有核种中如果不经过授粉,则在花后1～3日内子房中促进物质的水平即迅速降低而抑制物质水平则仍然保持很高。授粉引起了子房中促进物质水平的迅速增高与抑制物质水平的显著降低。天然单性结果品种与有核品种的主要区别在于开花后1～3天内前者的子房中在不授粉情况下也具有较高的促进物活性,而抑制物活性则大为降低。人工用外源生长物质诱发南瓜单性结果时,也同样引起内源促进物的急增而抑制物则完全消失。 这些结果说明了授粉受精的作用不仅在于引起子房中生长促进物的活化,而且也消除或对抗了生长抑制物的作用。座果与果实的继续发育受到生长促进物与抑制物间一定平衡状态的控制。本文对生长抑制物质的生物学意义也进行了讨论。     

红松种子中过氧化物酶活性与萌发的关系以及种子中的代谢抑制物

层积处理和去除种皮处理对红松种子萌发能力及过氧化物酶的活性有明显的影响。过氧化物酶在种子的休眠机理中可能有一定意义。红松种子的粗提物除能抑制种子萌发外,还有强烈抑制过氧化物酶活性和呼吸的能力。通过纸层析和薄层层析初步鉴定出,这些物质与抑制萌发的物质并非同一物质。粗提物中至少有两种过氧化物酶的抑制物,它们具有不同的稳定性。对酶活性抑制的时间曲线有不同影响,对不同同工酶和不同生物来源的过氧化物酶表现出不同的抑制。     

贵州石笔木种子内源有机化合物及对种子萌发的影响

该研究采用浸提法和GC-MS检测了贵州石笔木种子内源有机化合物种类、相对含量,以及种皮和胚乳在不同溶剂、温度和浓度条件下的浸提物活性。结果表明:(1)种皮和胚乳中皆含有有机酸、烯、酯、醇、醛、酚6类相对含量较高的有机化合物,其中种皮含有机酸7种、烯类1种、酯类5种、醇类3种、醛类3种、酚类1种,胚乳含有机酸6种、烯类1种、酯类1种、醇类1种、醛类1种、酚类1种。(2)种皮浸提物活性显著高于胚乳浸提物活性(P0.05),且其烯、醛、醇和酯类含量分别高出胚乳含量的8.78%、2.66%、2.15%和1.70%,可能是对种子萌发起主要作用的内源抑制物质。(3)不同条件下处理的浸提液均能显著抑制白菜种子发芽及幼苗生长,浸提液抑制物活性表现为醇溶剂大于水溶剂,并随着浸提液浓度的升高而增大、随着浸提温度的升高而增强,在初始温度为100℃时,浸提液抑制活性达到最大值。贵州石笔木种子的内源有机化合物在种子萌发过程中发挥着不同程度的抑制作用,探索其与种子萌发的作用机制,解决种子萌发育苗的关键技术及在农林业生产中应用,这在植物种子的生物学特性及萌发生理研究方面具有重要意义。     

珙桐种子休眠原因研究初报

珙桐内果皮和种子的水浸提液及乙醚提取液中均含有抑制物质;种子中的抑制物活性主要分布在被发达的胚乳包裹着的子叶内。层积一年后,内果皮及种子中的抑制物活性并未完全消失,但已明显减弱。珙桐种胚的形态后熟以胚芽在层积过程中完成形态分化为主要特征。     

柿树开花期子房中生长抑制物质的鉴定及其与子房发育的关系

从开花期柿树的未授粉子房中提取并分离出生长抑制物质,用紫外吸收光谱与显色反应法鉴定出抑制物的主要成分是脱落酸(ABA)和水杨酸。利用茄子子房试验测定了这两种抑制物对子房发育的影响,并与标准ABA、水杨酸及其它酚类化合物进行比较。实验的结果表明:ABA、水杨酸与提取抑制物都显著地抑制了授粉子房的发育,其中提取抑制物的抑制作用又比单用ABA或单用水杨酸者更为强烈。其他酚类如肉桂酸、咖啡酸与0-香豆酸等对茄果发育只产生轻度的抑制作用。这些结果说明ABA与水杨酸可能是调节柿子子房发育的主要抑制物质。花期未授精的子房不能继续发育的原因,就是由于这些生长抑制物质累积的结果。     

人工混交林中杉木、桤木和刺楸细根养分迁移的初步研究

比较分析了杉木 桤木和杉木 刺楸混交林中杉木、桤木和刺楸活细根、死细根的N、P、K含量 .结果表明 ,桤木细根N迁移能力较强 ,刺楸较弱 ,杉木细根N不迁移 ;P在桤木和刺楸细根中迁移能力较强 ,而在杉木细根中基本不迁移 ;3个树种细根脱落前都将K迁移回树体内 .比较分析 2个混交林中活细根N、P、K在树种间的差异 ,发现在杉木 桤木混交林中桤木根部N可能向杉木迁移 ,而在杉木 刺楸混交林中刺楸根部K可能向杉木根部迁移 ,但迁移机制还有待于从根 土界面生态过程进行研究     

红松种子抑制物质的初步研究

本文研究了红松种子抑制物质的提取方法,并测定了抑制物质的总活性,以及分布在种子各部位(外种皮、内种皮、胚乳、胚)抑制物质的相对活性。从红松种子各部位提取的抑制物质,不仅对油菜种子萌发和油菜幼根生长有抑制作用,而且也对其层积后解除休眠变黄的红松离休胚有明显的抑制作用。 通过红松种子各部位抑制物质变化动态的初步测定,表明了红松种子的休眠可能与发芽的抑制物质存在有关系。干藏红松种子各部位提取物在层析谱上 R_f值为 0.6的区段都有明显的抑制作用。除此而外,外种皮提取物在R_f值0.7区段,胚提取物在R_f值0.4、0.9区段也有明显的抑制作用。但干藏种子经过层积之后(即层积种子),各部位提取物与上述区段R_f值相比,其抑制作用有明显的下降。尤其是层积红松种子外种皮抑制物质基本消失或显著减少。 抑制物质常温下易溶于 95％乙醇、丙酮、水,其次是甲醇和乙醚,难溶于氯仿,不溶于苯。对高温(100℃)较稳定。     

A Preliminary Study on Neutral and Acid Inhibitors in the Seed of Kalopanax scptemlobus

The method of extraction, isolation, purification and identification of the neutral and acid inhibitors in the seeds of Kalopanax Septemlobus was given. It is proved that the neutral inhibitor is coumarin, the acid inhibitor is abscisic acid (ABA) by means of paper chromatograph, thin layer chromatograph, high performance liquid chromatograph (HPLC), color reaction and bioassay. The neutral inhibitor can strongly inhibite not only the seed germination of Brassica Chinensis and the radical elongation, but also that of the seed of Kalopanax septemlobus. The study showed that ABA and coumarin exist in the seed coat, endosperm and pericarp of the Kalopanax Septemlobus seed. Both ABA and coumarin can transport from seed cover (pericarp, seed coat)to the interior (endosperm, embryo) as the seeds were stored in refrigerator. In addition, the different results of extracting neutral inhibitor with water, methanol, and alcohol were compared in this paper.     

Abscisic acid in dormant apple seed tissues - a rapid purification scheme using pre-packed columns and GCMS-SIM quantitation

A rapid small scale procedure has been developed for quantitative abscisic acid (ABA) determination by gas chromatography - mass spectrometry with selected ion monitoping (GCMS-SIM). Extracts of apple seeds ( Malus domestica cv. Northern Spy) were passed first through 3 ml C₁₈ columns, and then through 3 ml silica gel columns. GCMS-SIM quantitation of ABA was done by adding an internal standard, hexadeuterated ABA. Ten sample extracts could be purified and analyzed by GCMS in 5 to 6 h, offering a quick and precise method to laboratories equipped with GC-MS instrumentation. The endosperm membrane and seed coat of apple seeds contained 2.5-3 times the ABA concentration found in the embryonic axis and cotyledons, supporting the hypothesis that it may be the ABA content of the membrane and seed coat that explains their inhibitory qualities with respect to seed germination.     

Endosperm-limited Brassicaceae seed germination: abscisic acid inhibits embryo-induced endosperm weakening of Lepidium sativum (cress) and endosperm rupture of cress and Arabidopsis thaliana

The endosperm is a barrier for radicle protrusion of many angiosperm seeds. Rupture of the testa (seed coat) and rupture of the endosperm are two sequential events during the germination of Lepidium sativum L. and Arabidopsis thaliana (L.) Heyhn. Abscisic acid (ABA) specifically inhibits the endosperm rupture of these two closely related Brassicaceae species. Lepidium seeds are large enough to allow the direct measurement of endosperm weakening by the puncture force method. We found that the endosperm weakens prior to endosperm rupture and that ABA delays the onset and decreases the rate of this weakening process in a dose-dependent manner. An early embryo signal is required and sufficient to induce endosperm weakening, which afterwards appears to be an organ-autonomous process. Gibberellins can replace this embryo signal; de novo gibberellin biosynthesis occurs in the endosperm and weakening is regulated by the gibberellin/ABA ratio. Our results suggest that the control of radicle protrusion during the germination of Brassicaceae seeds is mediated, at least in part, by endosperm weakening. We propose that Lepidium is an emerging Brassicaceae model system for endosperm weakening and that the complementary advantages of Lepidium and Arabidopsis can be used in parallel experiments to investigate the molecular mechanisms of endosperm weakening.     

Metabolism of carbohydrates during the development of seeds of the brazilian rubber tree [Hevea brasiliensis (Willd. Ex Adr. de Juss) Muell.-Arg.]

This work aimed at the assessment of the metabolism of carbohydrate during the development of the seeds of Brazilian rubber trees. The enzymatic activity of the acid invertase, neutral invertase and sucrose synthase (SuSy) and the levels of total soluble sugars (TSS), reducing sugars (RS) and sucrose were evaluated separately in each part of the fruit and seed—pericarp, seed coat, embryo and endosperm—on different days after the pollination (DAP). Based on the results obtained in this study, it is possible to conclude that in the beginning of the development of the rubber tree seeds, until 95 DAP, the endosperm presents high concentration of RS and low concentration of sucrose. After this period, the endosperm of the seed initiates starch accumulation and the concentration of RS decreases followed by the increase in the concentration of sucrose, presenting, after 120 DAP, an inversion of concentration of these two sugars. In the embryo, the levels of TSS, RS and sucrose show significant increase with the progress of the seed development. In the endosperm, the transition of the division phase and cell expansion for the storage of reserve material seem to occur around 120 DAP and is to be controlled mainly by the enzymes acid invertase and SuSy, while in the embryo, such transition seems to occur around 135 DAP and is to be controlled mainly by the enzymes acid and neutral invertases.     

The promoter of the asi gene directs expression in the maternal tissues of the seed in transgenic barley

The bifunctional alpha-amylase/subtilisin inhibitor (BASI) is an abundant protein in barley seeds, proposed to play multiple and apparently diverse roles in regulation of starch hydrolysis and in seed defence against pathogens. In the Triticeae, the protein has evolved the ability to specifically inhibit the main group of alpha-amylases expressed during germination of barley and encoded by the amyl gene family found only in the Triticeae. The expression of the asi gene that encodes BASI has been reported to be controlled by the hormones abscisic acid (ABA) and gibberellic acid (GA). Despite many studies at the gene and protein level, the function of this gene in the plant remains unclear. In this study, the 5'-flanking region (1033 bp, 1033-asi promoter) and the 3'-flanking region (655 bp) of the asi gene were isolated and characterised. The 1033-asi promoter sequence showed homology to a number of ciselements that play a role in ABA and GA regulated expression of other genes. With a green fluorescent protein gene (gfp) as reporter, the 1033-asi promoter was studied for spatial, temporal and hormonal control of gene expression. The 1033-asi promoter and its deletions direct transient gfp expression in the pericarp and at low levels in mature aleurone cells, and this expression is not regulated by ABA or GA. In transgenic barley plants, the 1033-asi promoter directed tissue-specific expression of the gfp gene in developing grain and germinating grain but not in roots or leaves. In developing grain, expression of gfp was observed specifically in the pericarp, the vascular tissue, the nucellar projection cells and the endosperm transfer cells and the hormones ABA or GA did not regulate this expression. In mature germinating grain gfp expression was observed in the embryo but not in aleurone or starchy endosperm. However, GA induced gfp expression in the aleurone of mature imbibed seeds from which the embryo had been removed. Expression in maternal rather than endosperm tissues of the grain suggests that earlier widespread assumptions that the protein is expressed largely in the endosperm may have been largely based on analysis of mixed grain tissues. This novel pattern of expression suggests that both activities of the protein may be primarily involved in seed defence in the peripheral tissues of the seed.     

Abscisic acid and osmoticum prevent germination of developing alfalfa embryos,but only osmoticum maintains the synthesis of developmental proteins

Developing seeds of alfalfa (Medicago sativa L.) acquire the ability to germinate during the latter stages of development, the maturation drying phase. Isolated embryos placed on Murashige and Skoog medium germinate well during early and late development, but poorly during mid-development; however, when placed on water they germinate well only during the latter stage of development. Germination of isolated embryos is very slow and poor when they are incubated in the presence of surrounding seed structures (the endosperm or seed coat) taken from the mid-development stages. This inhibitory effect is also achieved by incubating embryos in 10^?5 M abscisic acid (ABA). Endogenous ABA attains a high level during mid-development, especially in the endosperm. Seeds developing in pods treated with fluridone (1-methyl-3-phenyl-5[3-(trifluoromethyl)-phenyl]-4(1H)-pyridinone) contain low levels of ABA during mid-development, and the endosperm and seed coat only weakly inhibit the germination of isolated embryos. However, intact seeds from fluridone-treated pods do not germinate viviparously, which is indicative that ABA alone is not responsible for maintaining seeds in a developing state. Application of osmoticum (e.g. 0.35 M sucrose) to isolated developing embryos prevents their germination. Also, in the developing seed in situ the osmotic potential is high. Thus internal levels of osmoticum may play a role in preventing germination of the embryo and maintaining development. Abscisic acid and osmoticum impart distinctly different metabolic responses on developing embryos, as demonstrated by their protein-synthetic capacity. Only in the presence of osmoticum do embryos synthesize proteins which are distinctly recognizable as those synthesized by developing embryos in situ, i.e. when inside the pod. Abscisic acid induces the synthesis of a few unique proteins, but these arise even in mature embryos treated with ABA. Thus while both osmoticum and ABA prevent precocious germination, their effects on the synthetic capacity of the developing embryo are quite distinct. Since seeds with low endogenous ABA do not germinate, osmotic regulation may be the more important of these two factors in controlling seed development.     

Abscisic Acid levels and seed dormancy

Dormant seeds from Fraxinus species require cold-temperature after-ripening prior to germination. Earlier, we found that abscisic acid (ABA) will inhibit germination of excised nondormant embryos and that this can be reversed with a combination of gibberellic acid and kinetin. Using Milborrow's quantitative “racemate dilution” method the ABA concentration in 3 types of Fraxinus seed and pericarp were determined. While ABA was present in all tissues, the highest concentration was found in the seed and pericarp of dormant F. americana. During the chilling treatment of F. americana the ABA levels decreased 37% in the pericarp and 68% in the seed. The ABA concentration of the seed of the nondormant species, F. ornus, is as low as that found in F. americana seeds after cold treatment. Experiments with exogenously added ABA solutions indicate that it is unlikely that the ABA in the pericarp functions in the regulation of seed dormancy. However, the ABA in the seed does seem to have a regulatory role in germination.     

Proanthocyanidins Inhibit Seed Germination by Maintaining a High Level of Abscisic Acid in Arabidopsis thaliana

Proanthocyanidins (PAs) are the main products of the flavonoid biosynthetic pathway in seeds, but their biological function during seed germination is still unclear. We observed that seed germination is delayed with the increase of exogenous PA concentration in Arabidopsis. A similar inhibitory effect occurred in peeled Brassica napus seeds, which was observed by measuring radicle elongation. Using abscisic acid (ABA), a biosynthetic and metabolic inhibitor, and gene expression analysis by real-time polymerase chain reaction, we found that the inhibitory effect of PAs on seed germination is due to their promotion of ABA via de novo biogenesis, rather than by any inhibition of its degradation. Consistent with the relationship between PA content and ABA accumulation in seeds, PA-deficient mutants maintain a lower level of ABA compared with wild-types during germination. Our data suggest that PA distribution in the seed coat can act as a doorkeeper to seed germination. PA regulation of seed germination is mediated by the ABA signaling pathway.