Regulatory system of gene expression by novel artificial anti-gene chemicals in plants

植物中新型人工抗基因化学物质的基因表达调控系统

• 批准号: 11450316
• 负责人: YOSHIDA Kazuya
• 金额: $ 9.34万
• 依托单位: NARA INSTITUTE OF SCIENCE AND TECHNOLOGY
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (B)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2001
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11450316/
• 关键词: antisense method; plant cells; transgenic plant; PAL box sequence; peroxidase gene; promoter; transcription factor; アンチセンス化合; phosphorothioate; タバコ培養細胞; 熱ショックプロモーター; レポーターGUS遺伝子

[aim] Development of suppression method of gene function by anti-gene chemicals (S-oligo).[experiments]1. Movement of S-oligo in plant cells.2. Suppression of reporter gene function by S-oligo.3. Suppression of transcription factor gene by anti-sense RNA method.[Results and discussion]1. Total DNA was isolated from cultured tobacco cells (BY2) that were cultured in liquid medium containing S-oligo. It was confirmed with Southern blotting analysis and confocal laser scanning microscopy analysis that S-oligo was existence in the BY2 cells.2. We could not detect tie suppression of gene function by S-oligo. Quantitative analysis with in vitro translation system is necessary as next experiment.3. We have isolated a tobacco Ntlim1, as a trans factor binding to a PAL-box motif of the prxC2 promoter. We found that antisense expression of NtUm1 in transgenic plants carrying the prxC2 promoter : GUS chimeric construct decreased not only tie level of tie basal and the wounded-induced expression of tie GUS reporter gene but also the extent of tie wound indudbility of tie prxC2 promoter itself. This result indicates that Ntlim1 is required for the basal level of prxC2 promoter activity as well as its upregulation under wound stress.

[目的]建立抗基因药物抑制基因功能的方法(S-寡核苷酸)。[实验]1.S寡核苷酸在植物细胞中的运动。S寡核苷酸抑制报告基因功能。用反义核糖核酸方法抑制转录因子基因。[结果与讨论]1.从含S寡核苷酸的液体培养烟草细胞(BY2)中提取总DNA。经Southern印迹分析和激光共聚焦扫描显微镜分析证实，BY-2细胞中存在S寡核苷酸。我们没有检测到S寡核苷酸对基因功能的抑制。体外翻译系统的定量分析是下一步实验的必要手段。我们已经分离到一个烟草Ntlim1，作为与prxC2启动子的PAL盒基序结合的反式因子。我们发现，在携带prxC2启动子：GUS嵌合结构的转基因植株中，NtUm1的反义表达不仅降低了Tie碱基的TiE水平和创伤诱导的TiGUS报告基因的表达，而且还降低了Tie PrxC2启动子本身的结合程度。这一结果表明，Ntlim1是PrxC2启动子活性基础水平所必需的，也是创伤应激下PrxC2启动子活性上调所必需的。

项目成果

Phlla Kaothien et al.: "A cis-element containing PAL-box functions in the expression of the wound-inducible peroxidase gene of horseradish"Plant Cell Report. 19巻. 558-562 (2000)

Phlla Kaothien 等人：“含有 PAL 盒的顺式元件在辣根伤口诱导过氧化物酶基因的表达中起作用”植物细胞报告 19. 558-562 (2000)。

Kaothien, P., Shimokawatoko, Y., Kawaoka, A., Yoshida, K., Shinmyo, A.: "A cis-element containing PAL-box functions in the expression of the wound-inducible peroxidase gene of horseradish"Plant Cell Reports. 19巻. 558-562 (2000)

Kaothien，P.，Shimokawatoko，Y.，Kawaoka，A.，Yoshida，K.，Shimyo，A.：“含有 PAL 盒的顺式元件在辣根伤口诱导过氧化物酶基因的表达中发挥作用”植物细胞报告。第 19 卷。558-562 (2000)

Kaothien, P.,m Kawaoka, A., Ebinuma, H., Yoshida, K., and Shinmyho, A.: "Ntlim1, a PAL-box binding factor, controls the promoter activity of horseradish wound-induced peroxidase gene"Plant Mol. Biol.. (in press). (2002)

Kaothien, P.,m Kawaoka, A.、Ebinuma, H.、Yoshida, K. 和 Shinmyho, A.：“Ntlim1，一种 PAL-box 结合因子，控制辣根伤口诱导的过氧化物酶基因的启动子活性”植物

Kaothien, P., Kawaoka, A., Ebinuma, H., Yoshida, K., Shinmyo, A.: "Ntliml, a PAL-box binding factor, controls promoter activity of the horseradish wound-inducible peroxidase gene"Plant Molecular Biology. (印刷中). (2002)

Kaothien, P.、Kawaoka, A.、Ebinuma, H.、Yoshida, K.、Shinmyo, A.：“Ntliml，一种 PAL-box 结合因子，控制辣根伤口诱导过氧化物酶基因的启动子活性”植物分子生物学（正在出版）。

新名惇彦, 吉田和哉: "植物代謝工学ハンドブック"エヌ・ティー・エス(印刷中). (2002)

Atsuhiko Niina、Kazuya Yoshida：“植物代谢工程手册”NTS（印刷中）。