Symbiotic engineering with green manure leguminous plant and nitrogen-fixing rhizobia

绿肥豆科植物与固氮根瘤菌共生工程

• 批准号: 08455380
• 负责人: MUROOKA Yoshikatsu
• 金额: $ 4.67万
• 依托单位: Osaka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1996
• 资助国家: 日本
• 起止时间: 1996 至 1998
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-08455380/
• 关键词: nitrogen-fixation; nodule; Mesorhizobium huakuii; symbiont; leguminous plant; Astragallus sinicus; symbiotic engineering

1) Physical map of a large symbiotic plasmid in Mesorhizobium huakuii subsp. rengei :A large plasmid, pRhYM was prepared from M.huakuii subsp. rengei B3, which was isolated from renge-sou (Astragalus sinicus), by pulse field electrophoresis. The gene libraries were constructed from DNA fragments of the plasmid with a super cosmid vector. The libraries were analyzed by hybridization and made contig of about 300 kb.2) Nucleotide sequence of genes involved in nitrogen fixation and nodulation :A total about 80 kb nucleotide sequence of the symbiotic plasmid pRhYM was determined. On this sequence, several clusters of the nif and nod genes were located.3) Development of transformation system of M.huakuii :By using pBBR plasmid vector, M.huakuji subsp. rengei was transformed at the efficiency of 10^5.4) Nodule formation on renge-sou with the transgenic Rhizobium :The transformed strain of M.huakuii subsp. rengei was used to infect renge-sou seeds. The roots formed nodules, and the plasmid pBBR was isolated from the nodules.Thus, development of symbiotic engineering is now progress both in renge-sou plant and Rhizobium as a model.

1)华癸中慢生根瘤菌大共生质粒的物理图谱。rengei：从华癸支原体（M.huakuii subsp.）Rengei B_3是从紫云英（Astragalus sinicus）中分离得到的一种多糖。用超粘粒载体从质粒的DNA片段构建基因文库。2）固氮和固氮相关基因的核苷酸序列测定：共测定了共生质粒pRhYM约80 kb的核苷酸序列。3）华癸木霉转化体系的建立：以pBBR质粒载体为载体，将华癸木霉亚属（M.huakuji subsp.）4）用转基因根瘤菌对人参进行转化，转化后的菌株华癸根瘤菌（M.huakuii subsp.）使用Rengei感染Renge-sou种子。从根瘤中分离出质粒pBBR，以根瘤菌为模式菌，以人参植株为研究对象，开展了共生工程研究。

项目成果

Y.Hashimoto: "Characterization of the hemB gene encoding delta-aminolevulinic acid dehydratase from Propionibacterium freudenreichii." J.Ferment.Bioeng.82. 93-100 (1996)

Y.Hashimoto：“来自费氏丙酸杆菌的编码 δ-氨基乙酰丙酸脱水酶的 hemB 基因的表征。”

Yoshikatsu Murooka: "Nodule specific novel genes induced by infection of Rhizobium huakuii bv. renge in Astragalus sinicus (Chinese milk vetch)." Proceedings of the 11^<th> International Congress on Nitrogen Fixation, Paris, France, July 20-26,1997.339 (1

Yoshikatsu Murooka：“紫云英中的 Rhizobium huakuii bv. renge 感染诱导的根瘤特异性新基因。”

Yoshikatsu Murooka: "Nodule specific novel genes induced by infection of Rhizobium huakuii bv.renge in Astragalus sinicus (Chinese milk vetch)" Proceedings of the 11th International Congress on Nitrogen Fixation. 339- (1998)

Yoshikatsu Murooka：“紫云英中感染根瘤菌 huakuii bv.renge 诱导的根瘤特异性新基因”第 11 届国际固氮大会论文集。

Yoshikatsu Murooka: "Recent advances in studies on Streptomyces cholesterol oxidase and gene clusters involved in steroid catabolism in Prokaryotes." Actinomycetology. 10. 1-11 (1996)

Yoshikatsu Murooka：“链霉菌胆固醇氧化酶和原核生物类固醇分解代谢相关基因簇的研究最新进展。”

Yoshikatsu Murooka: "The monoamine regulon including synthesis of Arylsufatase and monoamine oxidase in bacteria." Bioscience Biotechnology Biochemistry. 60. 935-941 (1996)

Yoshikatsu Murooka：“单胺调节子包括细菌中芳基硫酸酯酶和单胺氧化酶的合成。”