The quantitative evaluation offixed nitrogen by the endophytic bacteria in sweet potato and sugarcane

植物体内的固氮细菌被称为植物内生固氮细菌。固氮植物仅限于与根瘤菌有共生关系的豆科植物和与Frankia有共生关系的近十种植物。从巴西甘蔗中分离到固氮内生细菌，其固氮率可达60-70%，在甘蔗中，固氮内生细菌栖息。固氮对植物总氮的贡献率与根瘤菌-豆科植物共生的贡献率相当。固氮内生细菌主要从湿地水稻、小麦、玉米、高粱和各种牧草中分离得到。这表明非豆科植物也具有固氮功能。但是，也存在一些问题， ...更多信息 解决实现这一点。这两个步骤是筛选出有益的内生细菌并定殖到植物体内。本研究以平成16年至平成19年的4年时间为研究对象，探讨了内生固氮菌对植物的侵染、增殖以及内生固氮菌固氮量的估算方法。在研究期的前半期，用GFP标记的Herbaspirillum sp.进行了接种试验。并尝试从甘蔗和甘薯中分离出负责观察到的N2固定的内生固氮菌。此外，还通过接合和电穿孔技术获得了编码绿色荧光蛋白（GFP）的基因。研究后期，对分离自甘蔗和甘薯的内生固氮菌进行了接种效果和在非寄主植物中的内定殖能力的测定。同时，对甘蔗茎中固氮细菌的相对丰度进行了测定。NiF 8、Nil 5、N7和KTN 94 -88在甘薯茎、块茎、叶和根中的表达量最高。从上述材料中分离到39个菌株，其中甘蔗27个，甘薯12个。Herbaspirillum sp.，本实验中使用的肠杆菌属和泛菌属可以在其原始宿主以外的植物中建立，表明缺乏宿主特异性，尽管定殖程度可能取决于宿主和内生菌的组合而变化。本研究以甘蔗组培苗的根部为接种部位，主要研究了内生固氮细菌在甘蔗组培苗中的作用。但是，我们得出的结论是，将内生细菌导入甘蔗等无性繁殖植物的合适的传递方法是茎段真空渗透法。少

The nitrogen-fixing bacteria which inhabits the inside plant is being called nitrogen-fixation endophytic bacteria. The plant which fixes nitrogen is limited to the leguminous plants which have a symbiotic relationship with RhJzobia and about ten kinds of plants that have a symbiotic relationship with Frankia However, several twenty years ago ; the existence of the nitrogen-fixing endophitic bacteria was isolated from sugarcane in Brazil And also it was reported that the rate of fixed nitrogen to total nitrogen might reach 60-70% in the sugarcane that nitrogen- fixing endophytic bacteria inhabited. The rate of contribution of N2 fixation to total plant N is equal to the Rhizobium-legume symbiosis. As for the nitrogen fixing endophytic bacteria, they were isolated from wetland rice, wheat, maize, sorghum, and various forage grasses in addition to sugarcane. This showed the possibility that a nitrogen fixing function is given to the non-leguminous plants. But, there are some problems to … More solve to realize this. They were the selection of useful endophytic bacteria and a colonization to the inside plants. The infection and multiplication to the plant of the nitrogen fixing endophytic bacteria and the estimation method of the nitrogen fixed by endophytic diazotrophs was examined in this research.A research period is four years from the 16th year of Heisei until 19. At the first half of research period, the inoculation examination by GFP-labeled Herbaspirillum sp. was conducted. And the isolation of the endophytic diazotrophs, are responsible for the observed N2 fixation were tried from sugarcane and sweet potato. Moreover the gene encoding green fluorescent protein (GFP) via conjugation and electotroporation At the latter half of research period, inoculation effectiveness and internal colonization ability as an introduced in the non host plants of endophytic diazotrophs isolated from sugarcane and sweet potato was examined. And also the method for the amount of nitrogen fixed a period of duration was examined.As a result, we determined the relative abundance of N2-fixing enndophytic bacteria in the stem of sugarcane cvs. NiF8, Nil5, N7 and KTN94-88, and in the stem, tuber, leaf and root of sweet potato cvs. Koganesengan and Shiroyutaka and 39 isolates, 27 isolates from sugarcane and 12 isolates from sweet potato, were obtained from the above-mentioned materials. Herbaspirillum sp., Enterobacter sp. and Pantoea sp. used in this experiment could be established in plants other than their original host, suggesting a lack of host specificity, although the extent of colonization may vary depending on the combination of host and endophyte. And, higher bacterial numbers were detected in plant tissues inoculated with higher inoculum (10^8 cells/ml)concentration treatment.We have used the root part of the micropropagated sugarcane plant to the target mainly as an inoculation part of the useful N_2-fixing endophytic bacteria in this study. However we got the conclusion that a suitable delivery method for introducing endophytic bacteria into the vegetative propagation plants such as sugarcane is the vacuum infiltration method to stem pieces. Less