【作者】 黄有凯；

【导师】 蒋立科； 罗曼；

【作者基本信息】 安徽农业大学 ， 生物物理学， 2003， 硕士

【摘要】 人工合成肥料和农药对生存环境的污染，推进了人们对生态农业的认识，由此而产生的“崇尚自然”、“回归自然”已成为不可逆转的潮流。当前，一个以绿色食品(或有机农业产品)为中心的生产和消费热潮正在全球兴起。依国际农业生产标准规定：种植业禁止使用化学肥料和化学的农药，养殖业禁止使用抗生素和激素，产品贮存中禁止使用化学药品。这对于农业生产中发挥重要增产作用的植物生长促进物质无疑提出了更高的要求，世界有关研究者将注意力转向天然微生物肥料，由木霉产生的促进植物生长物质不仅能减轻大气污染，改善因长期使用化肥而使土壤结构破坏状况，而且能提高农作物品质，增进人类健康。 木霉(Trichoderma.SPP)属于半知菌类丛梗孢目，丛梗孢科真菌，(Gloiosporae)是一种普遍存在的真菌。在生物防治及促作物生长中有广阔的应用前景。木霉作为一种新的微生物肥料开发菌，其遗传学特性不稳定，促生长机制尚不明确。因此进一步选育出性能稳定的促作物生长的突变株，并研究其对植物生长各项生理生化指标的影响，具有重要的现实意义。 通过对哈茨木霉、绿色木霉和里氏木霉的发酵液浸种后的水稻、小麦萌发过程中的发芽率、发芽势、根长、芽长和根冠比的比较，发现50倍稀释的哈茨木霉发酵液对种子的萌发有较为显著的促进作用。但绿色木霉对种子的萌发的影响不明显，里氏木霉对种子的萌发有一定的抑制作用。据此，采用哈茨木霉作为出发菌株，通过离子束诱变，从大菌落株中筛选出促进生长的高产突变株——h-13菌株，通过对用该菌株发酵液处理后的水稻及小麦的种子的各项生理生化指标的测定，发现该菌株比其它菌株更能促进水稻和小麦的活力和发芽率，三天时小麦的总根长为3.03cm，比对照的1.42cm的两倍还多，并且能够明显提高种子发芽后的淀粉酶和蛋白酶的活力，其活力都是对照的两倍左右。用该发酵液喷施水稻幼苗，发现该菌株对水稻苗的株高、分蘖数及生物量都有较为明显的促进作用。从小菌落中筛选出营养缺陷突变株，发现营养缺陷突变株促植物生长不明显。采用正交实验设计，对突变株h-13菌株，在不同基质的培养基上的生长情况进行比较分析，筛选出以麸皮为碳源的最佳培养基，使五天时的产孢量是基础培养基的2.2倍。该菌株发酵液含有对水稻生长有显著促进作用的物质；经对喷施该菌株发酵液后的水稻硝酸还原酶（NR）活力及其N、P和K含量的测定，发现该菌株发酵液促进水稻生长与提高NR活力、增强对N、P和K的吸收有关。喷施该菌株发酵液可促使水稻叶片中的叶绿素A、B及总量的增加。叶绿素A、B比对照分别增加7石％～35刀％和11．5％～30二％，叶绿素总量的增幅为4刀％～17．8％。采用酶动力学扫描及垂直板凝胶电泳技术，对水稻喷施该菌株发酵液后，测定多酚氧化酶 OPOL过氧化物酶①OD）活性和同工酶带图谱的动态变化。与对照组相比，该菌株不仅能够提高P 和POD酶的活力，而且还能诱导产生新的同工酶谱带，但不同浓度发酵液对诱导合成的酶作用不同。稀释50倍的发酵液对POD活力有较大的阳性效应。高浓度的发酵液对即 活性有先抑制后出现提高的趋势，表明了该发酵液对叩 活性调节的程序性。另外，该菌株发酵液还能提高SOD酶的活力，诱导产生新的同工酶带谱。通过对大棚黄瓜大田试验，发现该菌株能够促进黄瓜幼苗的生长，其中50倍稀释后的发酵液喷施后的黄瓜比对照有更为明显的株高、根长、茎粗和鲜重。最后比对照增产23％。 综上所述，本研究通过绿色木霉、里氏木霉和哈茨木霉对植物的生长作用比较，以哈茨木霉为出发菌株，采用离子束注入的方法诱变筛选促植物生长高产菌株，然后以其发酵液研究其对小麦、水稻种子的发芽率、淀粉酶的影响，电镜扫描突变与非突变株的抱子形态的比较，分析过氧化物酶、多酚氧化酶和超歧物氧化酶的酶活力及同工酶、测氮、磷、钾含量研究其对植物生长的影响，阐叙其作用规律。与国内外同类研究比较，采用离子束注入筛选高效突变株，分析同工酶及氮、磷及钾的含量，有创新之处，本项研究若在小区域试验的基础上，进行大面积的田间实验，将有良好的开发前景。更多还原

【Abstract】 The pollution of synthesis fertilizer and pesticide to existence environmental makes people to recognize agriculture, an unreversion tide of "Adore Nature","Return Nature" has become. Now the tendency that green food acts as a center role in production and consumes has being grown. The international agricultural production criterion rule: forbid to use fertilizer and pesticide in the faming, forbid to use antibiotics and hormone in the cattle breeding, forbid to use chemical drug in the production saving. This gives a higher requirement to the substance of promoting plant growth, which plays an important role in the agricultural production. Many researchers have paid attention to natural microorganism manure. The substance of promoting plant growth from Trichoderma not only can relieve atmosphere pollution, ameliorate soil structure, which has been destroryed by fertilizer, but also can enhance quality of plant, rise human health.Trichoderma. SPP belong to cluster stem cohort of Deuteromycetes, Gloiosporae fungus. It is a kind of generally existent fungus. Trichoderma harziaiarum is not only a kind of biological control fungus that has been studied extensively, but also play a role for promoting plant growth. Trichoderma. SPP, as a new useful fungus of microorganism manure, its genetic peculiarity isn’t stability, the mechanism of promoting plant growth isn’t clear. It is important to screen a stable mutate strain of promoting plant growth and researching its effect on plant.Soaking seed of rice and wheat with zymolytic liquid of Trichoderma harziaiarum, Trichoderma DWC and Trichoderma viride, comparing with germinate ratio, germinate force, root length, bud length and ratio of root to bud, the results show that 50 times dilute zymolytic liquid of Trichoderma harziaiarum has a relatively notable promote role for germinate of seed .The effect of Trichoderma viride to germinate of seed is not evidence. Trichoderma DWC for seed sprout has certain inhibition. Trichoderma harziaiarum injected by ionic beam, guided to change, then screened a strain called h-13. In the zymolytic liquid of this strain there is a substance, which can promote germinate ratio of seed remarkably. Through determining every physiologicalbiochemical parameter of germinate seed of rice and wheat which deals with of muttagenic strains, the result shows that the strain of h-13 can promote germinate ratio and force more greatly than other strains. In 3 days, the totalled root length of wheat, which deals with zymolytic liquid of H-13, is 3.03 cm; it is about two times than the 1.42 cm of contrast. For the more, It can raise vigour of amylase and protease in germinate seed obviously, and its activity is about the two times than compares controls. Trichoderma harziaiarum can resist disease of plant, and promote plant growth. So it is very useful to screen a kind of culture medium, which is both of high efficient and cheap. Anglicizing and comparing the growth of Trichoderma harziarum in different culture medium with orthogonal design experiment, we got the best culture medium. The composition is: wheat bran 100g, FeSO4 100mg ,MgSO4 7H2O 1.25g ,K2HPO4 0.7g, KH2PO4 0.4g, NaNO3 7g. The muttagenich-13 strain, which was injected by ionic beam and screened from Trichoderma harziaiarum, can promote the growth of rice remarkably. In the experiment, we use zymolytic liquid of h-13 strain sprinkle on the seedling of rice, and determine nitrate reductase activity and N, P, K content. The results suggest that the effect of promoting plant growth of the strain has a relation to the enhance of nitrate reductase activity and the increase of N, P and K absorption. Zymolytic liquid of Trichoderma harziaiarum can increase the content of chlorophyyll B and A in the leaf of rice . Chlorophyyll A and B increase 7.6% ~ 35.0% and 11.5% ~ 30.2% than compares controls. The total of chlorophyyll amount increase is about 4.9% ~ 17.8%. Trichoderma harziaiarum plays a role of antagonic ability to pathogenic microbes and of inducing plant to produce a resistivity of dis更多还原