| Research Abstract |
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
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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
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