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

植物体内的固氮菌被称为固氮内生细菌。固氮植物仅限于与RhJzobia有共生关系的豆科植物和与Frankia有共生关系的约10种植物；固氮内生细菌是从巴西甘蔗中分离出来的，据报道，在固氮内生细菌所栖息的甘蔗中，固氮占总氮的比例可达60-70%。氮素固定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。除了甘蔗外，从湿地水稻、小麦、玉米、高粱和各种牧草中也分离到了固氮内生细菌。这说明了非豆科植物具有固氮功能的可能性。但是，…也有一些问题更多的解决方案来实现这一点。它们是对有用的内生细菌的选择和对植物内部的定植。本研究对固氮内生细菌对植物的侵染繁殖及固氮菌固氮的估算方法进行了研究。研究周期为平成16年至19年，为期4年。在研究的前半段，对GFP标记的草本菌进行接种检测。进行了一次。并从甘蔗和甘薯中分离出内生固氮菌，对观察到的氮素进行了固氮试验。此外，对研究后期接合和电转化的绿色荧光蛋白(GFP)基因在甘蔗和甘薯内生固氮菌的非寄主植物中的接种效果和内定殖能力进行了检测。并对施氮量固定时间段的方法进行了研究，测定了甘蔗CVS茎中固氮菌的相对丰度。NiF8、Nil5、N7和KTN94-88，以及甘薯CVS的茎、块茎、叶和根中。从上述材料中分离到Koganesengan和Shiroyutaka菌株39个，其中甘蔗菌株27个，甘薯菌株12个。草杆菌属、肠杆菌属。和Pantoea sp.在本实验中使用的植物可以在其原始寄主以外的植物中建立，这表明缺乏寄主特异性，尽管定殖的程度可能因寄主和内生真菌的组合而异。接种浓度较高(108个/ml)的甘蔗植株组织中细菌数量较高。本研究以甘蔗微繁植株的根部为靶标，主要作为有效固氮内生细菌的接种部分。然而，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输送方法是真空渗滤法。较少

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