2003 Fiscal Year Final Research Report Summary
Studies on developments of cell and tissue culture
| Project/Area Number |
13460065
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| Research Category |
Grant-in-Aid for Scientific Research (B)
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| Allocation Type | Single-year Grants |
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| Section | 一般 |
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| Research Field |
林学
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| Research Institution | Tokyo University of Agriculture and Technology |
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Principal Investigator |
KUBO Takafumi Tokyo University Agriculture And Technology, Graduate School of Agriculture, Professor, 大学院・農学研究科, 教授 (00015091)
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| Co-Investigator(Kenkyū-buntansha) |
OGITA Shinjiro Toyama Prefectural University, Faculty of Technology, Assistant Professor, 工学部, 助手 (50363875)
SASAMOTO Hamako Yokohama National University, Environmental and Information Science, Professor, 環境情報研究院, 教授 (60334629)
KAWAI Shinya Tokyo University Agriculture And Technology, Faculty of Agriculture, Associate Professor, 農学部, 助教授 (90202027)
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| Project Period (FY) |
2001 – 2003
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| Keywords | Embryogenic cells / Pinus densiflora / cryptomeria japonica / Somatic embryo / Amino acid / Protoplast culture / Micro-plate assay method / CjNdIy |
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| Research Abstract |
(1)In subculture of embryogenic cells (ECs) from immature embryo of Pinus densiflora, more effevtve culture conditions were mDCR medimun which added L-glutamine of 1250mg/l and polyvinyl pyrrolidone(PVP) of 1000mg/l, resulting in higher proliferation of ECs with good quality. Furthermore, in formation of somatic embryo from ECs, additions of abscisic acid (ABA) of 30μM, maltose of 10% and polyethylene glycols (PEG) of molecular weight 8000 were effective in the development.
(2)In Larix leptolepis, Picea jezoensis and Crypbomeria japonica, characteristics of the culture cells were clarified b**** the morphological observation and a quantitative analysis of endogenous amino acid, using micro-HPLC Consequently, effect of time amino acid was confirmed in the subculture of ECs, and an addition of a certain concentration of amino acid promoted development of ECs with clear and large embryo region and long suspensors. Thus result suggests possibility of somatic embryo formation. In recalcitran
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t species, like Cryptomeria japonica, it is considered that protocols of the analysis and evaluation by monitoring endogenous amino acid contents are effective for selecting the cells with high differential ability in the early stage, which result finally in somatic embryo
(3)A suitable condition for isolation of protoplast and an optimum combination of medium and plant hormone for proliferation of protoplast generally vary according to plant species. Micro-plate (24 well) assay method that had been. established for protoplast culture of Populus alba, is adaptable to investigate culture conditions of coniferous tree such as L.leptolepis and C.japonica. By culturing small amount of protoplasts in a well of 96 well micro-plate, an optimal kind and concentration of plant hormone(s), and/or cell density was easily determined.
(4)We determined the entire nucleotide sequence of GjNdly with the exception of the 5'-untranslationed promoter region. Although no proline-rich region has previously been detected in the gene product from gymnosperms, we found such a region in the amino-terminal region of the deduced amino acid sequence from Cryptomeria japonica. Less
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