Application of no utility plant resources by a recombinant DNA technology
Grant-in-Aid for Scientific Research (B).
|Allocation Type||Single-year Grants|
|Research Institution||NARA INSTITUTE OF SCIENCE AND TECHNOLOGY|
SHINMYO Atsuhiko Nara Institute of Science and Technology, Graduate School of Biological Sciences, Professor, バイオサイエンス研究科, 教授 (30029235)
KATO Ko Nara Institute of Science and Technology, Graduate School of Biological Sciences, Assistant Professor, バイオサイエンス研究科, 助手 (80283935)
SEKINE Masami Nara Institute of Science and Technology, Graduate School of Biological Sciences, Assistant Professor, バイオサイエンス研究科, 助手 (70226653)
YOSHIDA Kazuya Nara Institute of Science and Technology, Graduate School of Biological Sciences, Associate Professor, バイオサイエンス研究科, 助教授 (50252622)
|Project Period (FY)
1998 – 2000
Completed(Fiscal Year 2000)
|Budget Amount *help
¥12,900,000 (Direct Cost : ¥12,900,000)
Fiscal Year 2000 : ¥3,100,000 (Direct Cost : ¥3,100,000)
Fiscal Year 1999 : ¥4,000,000 (Direct Cost : ¥4,000,000)
Fiscal Year 1998 : ¥5,800,000 (Direct Cost : ¥5,800,000)
|Keywords||sweet potato / heat shoch protein gene / promoter / artificial regulation of gene expression / hormone-like compound / peroxydase gene / ペルオキシダーゼ / シロイヌナズナ / 熱ショック遺伝子 / 転写因子 / 位置効果 / タバコ細胞 / アルコールデヒドロゲナーゼ遺伝子 / 植物バイオマス / 廃棄物 / 西洋ワサビ / 誘導型プロモーター|
The project focus on the accumulation of valuable proteins in leaves of rootcrop, which have photosynthetic and metabolic activities even at the harvesting, by a recombinant DNA technology. Target plant and enzyme were sweat potato, Ipomoea batatas as a typical rootcrop popular in Japan and horseradish peroxidase as a useful plant origin enzyme. Development of artificial gene expression system is an important basic technology in this purpose. The following results were obtained :
1) Introduction and expression of horseradish peroxidase genes, prxC1a andprxC2 in sweat potato.
The cDNA of prxC1a or prxC2 gene was ligated downstream of CaMV35S promoter and introduced to callus of sweat potato together with hygromycin-resistant gene using Agrobacterium. Several transgenic plants resistance to hygromycin showed significant accumulation of mRNA, protein and enzyme activity of the peroxidase.
2) Construction of inducible gene expression system.
To accumulate high amount of useful proteins in swea
t potato leaves without damage of basic metabolic activity, inducible system of transgenes at suitable developing stage is required. One of approach is application of heat-inducible promoter of Arabidopsis thaliana HSP18.2 gene. The HSP18.2 promoter was ligated to GUS reporter gene and introduced to tobacco cultured cells (Nicotinana tabacum BY2). When incubation temperature was sifted from 27℃to 35℃, GUS activity increased 1,000 times after 2 hr.Modification of this system will be useful for our goal.
Biosynthesis of virginiamycin is induced by vinginia butanolides (VB) in Streptomyces virginiae. The mechanism of induction was elucidated already. S.vinginiae synthesizes constitutively repressor protein and the repressor binds to operator sequence upstream a coding region of virginiamycin biosynthetic enzymes. Repression is released by binding of VB to the repressor. Therefore, a structure gene of the repressor was driven by CaMV35s promoter in tobacco BY2 cells, and accumulation of the repressor protein was detected. The operator sequence was placed in various positions upstream GUS structure gene, and introduced to nudear chromosome of BY2 cells harboring the repressor gene. Addition of VB in culture medium of the BY2 cells induced GUS activity significantly. Optinization of number of the operator sequence and position to be introduced upstream of the GUS gene was established. The system will be used to practical production of peroxidase in sweat potato by spraying of VB on the leaves after development of potato tuber. Less
Research Output (1results)