【作者】 张勇；

【导师】 曾明；

【作者基本信息】 西南农业大学 ， 果树学， 2003， 硕士

【摘要】 自然条件下，绝大多数植物都可形成菌根，其中丛枝菌根(Arbuscular Mycorrhiza，AM)是最普遍的类型。AM是植物根系和丛枝菌根真菌(Arbuscular Mycorrhizal Fungi，AMF)形成的共生体(Symbiont)，目前已经确知，AM在促进植物生长，提高养分吸收率，生态系统养分循环及保护植物抵御不良环境胁迫中起关键作用，因此人们对AM生物技术及其在生产实践中的应用进行了广泛研究，取得了较大进展。近几年，国内外学者在AMF生长促进物质及AMF离体培养方面都进行了一些积极的探索，但目前为止，AMF还不能在离体培养条件下完成其生命周期，种苗繁育以及研究工作中应用的AMF菌剂仍是已形成AM的宿主植物根系或带AMF的根系土壤，给研究和实际应用带来诸多不便。 本论文从AMF应用基础研究出发，以摩西球囊霉(Glomus mosseae)、珠状巨孢囊霉(Gigaspora margarita)为主要试材，从多胺(Polyamine，PA)及银杏根系甲醇溶提物对离体条件下AMF生长发育的影响、AMF对葡萄微繁苗的侵染过程及PA对菌根化葡萄微繁苗生长发育的影响等方面进行了如下试验研究： 试验1 PA对离体条件下AMF生长效应的研究 PA是一类具有生理活性的低分子量含氮碱，被认为是一种“第二信使”，可与DNA、RNA结合，影响核酸和蛋白质的代谢，明显促进细胞分化，对植物的生长发育具重要的调控作用。试验1研究离体培养条件下PA(腐胺：Putrescine，PUT；亚精胺：Spermidium，SPD；精胺：Spermine，SPM)及PA生物合成抑制剂(甲基乙二基-双(脒基腙)：Methylglyoxal Bis(Guanylhydrazone)，MGBG)对AMF(Gl. mosseae，Gi. margarita)孢子萌发特性及菌丝生长 2003届项士学位论文：丛技菌根真菌离体培养及其对葡萄微繁苗生长效应的研究发育的影响。 试验结果表明，3种多胺类物质在 50ug／ffil200Ug／ffil浓度范围内，对 AMF生长发育具显著促进作用，而 500ug／nd浓度处理对 AMF生长发育表现强烈的抑制效应。MGBG（50Ug／tal－500ug／ml）对 AMF生长发育有较强的抑制作用，且可被外源 PA部分解除，但随浓度升高外源PA的恢复作用降低，5皿Ug／nd时无效。同时观察到肛M可诱导F抱于多芽管萌发，促进菌丝多分支的后继生长。作者认为AMF体内内源PA含量也许是其生长发育的限制因子。 试验发现，PA对AMF生长发育的影响具有两个特点：一是PA对离体培养条件下AMF泡于萌发的影响作用具有种间差异性，Gi．mamprita较 GL mossea。对 PA反应敏感；二是PA对AMF生长发育的促进作用具有二重性——低浓度促进，高浓度则有抑制作用。通常认为－NH。越多PA的生理活性越强，因而SPM活性最强，SPD次之，PUT最弱，这与本试验所观察的现象相符。另外，AMF继代菌丝对PA反应更敏感，但种间差异性不明显。 试验2＄杏根系甲醇溶提物对离体条件下AMF生长效应的研究 许多研究表明，植物根系提取物对AMF生长发育有一定影响，而宿主植物根系中黄酮类化合物与AMF的生长发育及AM共生体的形成密切相关。本试验研究银杏根系甲醇溶提物对离体培养条件下AMF生长发育的影响。 试验结果表明：银杏根系甲醇溶提物对离体条件下AMF（GI mosseae，Gi ma哪rita）生长发育有明显的促进作用，能显著提高抱子萌发率，增加菌丝生长量。与对照相比，浓度为20Oh－100％的甲醇溶提物均可显著促进 AMF的生长发育，80Orb甲醇溶提物效果最明显。银杏根系甲醇溶提物中含丰富的黄酮类化合物，溶提物中黄酮类物质的含量与甲醇洗脱剂的浓度有关，当浓度为80％时，黄酮含量最高。通过对银杏根系甲醇溶提物中黄酮类化合物含量与AMF生长发育相关性分析发现，溶提物中黄酮类化合物含量与离体条件下AMF生长发育表现为极显著的正相关关系：溶提物中黄酮含量越高，其对AMF生长发育促进作用越大。 试验3 ＊MF对葡萄（Ntik V切诈L）微繁苗生长效应的研究 葡萄是形成AM的植物，可被多种AMF侵染，且其生长发育在很大程度上依赖于AMF的侵染及发育良好的AM共生体的形成。目前，用离体培养技术快速繁育葡萄种苗己有很大发展，但葡萄微繁苗是在无菌条件下繁育的，缺少AMF的侵染，导致微繁苗生长势弱、抗逆性差、移栽成活率低。为此，对葡萄微繁茁及早接种适当的AMF，培育出菌根化葡萄微繁苗，提高微繁苗的移栽成活率，促进幼苗生长，可更加提高葡萄优良稀缺品种的繁育速度及数量。 试验 3研究了接种 AMF（GI mo．rseae）对离体条件下葡萄微繁苗增殖、生根及练苗时对葡萄微繁苗生长效应的影响。试验结果表明，AMF对葡萄微繁苗在离体培养及练苗培育阶段 2 摘要的生长发育均有重要的促进作用。AMF对葡萄外植体的增殖和生根有一定促进，尤其是增加了生根条数。试验发现，AMF能促进根系磷酸酶的活性，磷酸酶活性与AMF侵染率存在显著相关性。接种AMF可增强幼苗根系活力，促进根系对N、P等矿质养分的吸收和积累，并促进植株的光合作用，提高幼苗的生长势。 试验4PA对菌更多还原

【Abstract】 Mycorrhizae are symbiotic associations between plant roots and certain soil fungi and the almost plants can form the mycorrhiza structure in the natural condition. The arbuscular mycorrhiza (AM) is the commonest mycorrhizal association between host plant roots and seven genera of soil fungi (Arbuscular Mycorrhizal Fungi, AMF), which are classified to Eodogone , Glomus. Sclerocysfis. Acaulospora. Entrophospora. Gigaspora and Scutellospora. It was known that the AMF played a key role in nutrient cycling in the ecosystem and promoting plant growth and also protected host plants against environmental stress. Due to the positive effects of AMF, the studies and utilization of AM biotechnology were being paid enormous attention, and great advance was achieved all over the world. In recent years, researchers have made some progress in stimulative substances for AMF growth and development and AMF in vitro culture, but AMF couldn’t complete their life cycle in vitro. So, AMF inoculum used in commercial propagating plantlets or scientific researches are often host plant roots infected with AMF or rhizosphere soil with AMF, which brought many difficulties to the studies and utilization of AM biotechnology.With the two AMF (Glomus mosseae, Gigaspora margarita) being used as the main experimental materials, the experiments were carried out to study the effects of polyamine (PA) and methanol eluates fractionated from Ginkgo biloba root on the growth and development of AMF in vitro, the traits of AMF infection in grape (Vitis vinifera L.) root in vitro and the effects of PA on the growth and development of micropropagated grape plantlets as follows:Master Degree Dissertation, 2003 : Study on In vitro Culture of Arbuscular Mycorrhizal Fungi and TheirEffects on the Growth of Micropropagated Grape (Vitis vinifera L.)Experiment 1 Study on effects of PA on the growth and development of AMF in vitroPA are a sort of low molecular weight alkali with nitrogen element and have physiological activity. It can control the metabolism of nucleic acid and protein, promote cell differentiation and have important regulative effects on the plant growth by combined with deoxyribonucleic acid or ribonucleic acid. It was considered that PA was a kind of second messenger.hi experiment 1, the effects of PA (Putrescine, PUT; Spermine, SPM; Spermidium, SPD) and PA biosynthesis inhibitor (Methylglyoxal Bis (Guanylhydrazone), MGBG) on spore germination character and hyphal growth and development of AMF (Gl. mosseae, Gi. margarita) in vitro was studied. The results indicated that PUT, SPD and SPM had significant effects on spore germination rate and hyphal growth of AMF at the range of 50ug/ml~200ug/ml, but high concentration PA treatment (at 500ug/ml) indicated significantly inhibiting effect. From 50ug/ml to 500ug/ml, MGBG had negative effect on the growth and development of AMF and this effect could be partially relieved by exogenous application of PA, but this relief role was reduced when the concentration of MGBG increased. The author realized that the endogenous concentration of polyamine of AMF might be a limiting factor of the growth and development.The results showed that the effects of PA on the growth and development of AMF were dependent on the fungi species and the PA treatment concentration: the reaction of Gl. mosseae to PA was more sensitive than Gi. margarita^’, the effect of PA was positive in low concentration, but it changed into negative in high concentration. In generally, it was known that the more -NHa PA had, the more distinct physiological activity PA had. So, SPM had the most distinct activity, SPD had the secondly high activity and activity of PUT was vague, which coincided with the results obtained from this experiment.Experiment 2 Study on the effects of methanol eluates fractionated from Ginkgo biloba root on the growth and development of AMF in vitroSome researches have showed that eluates of plant root had some effects on the growth and development of AMF, at same time, the flavonoids of更多还原