Studies on the utilization of useful genes isolated from the wild plant for control of virus diseases

利用野生植物中分离的有用基因防治病毒病的研究

• 批准号: 11460024
• 负责人: OSAKI Takeshi
• 金额: $ 8.9万
• 依托单位: Osaka Prefecture University
• 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 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-11460024/
• 关键词: Wild plant; Cucumis figarei; Virus resistance; CF-RIP gene; Figaren; Antiviral protein; Control of virus disease; RNase

A cDNA coding a putative ribosome-inactivating protein(CF-RIP) from Cucunis figarei was cloned and sequenced. The entire cDNA contains an ORF encoding a protein of 286 amino acids which share conserved residues for ribosome-inactivating proteins(RIPS). CF-RIP share 54.8-64.5% identity with four RIPs from other cucurbits. RT-PCR analysis indicated that CF-RIP mRNA was expressed in leaf and stem tissues of Cucumis figarei.An antiviral protein, designated figaren, was purified from leaves of C.figarei and partially characterized. Column chromatography, SDS-PAGE and periodic acid-Schiff staining revealed that figaren is a glycoprotein with a molecular weight of 23 kDa. N-terminal amino acid sequencing indicated that figaren contained the conserved region for the S-allele-associated ribonucleases(RNases). In-gel RNase assay showed that figaren digested yeast RNAs. Fluorescence in situ lybridization revealed that figaren and Rnases(beef pancrease and RNase T1)at 1μ g/ml similarly inhibited C … More MV infection in cowpea leaves. Figaren and Rnases at 5-500ng/mlhad similar inhibitory effect on local lesion formation by CMV.CMV accumulation detected by ELISA in tobacco leaves treated with figaren 6 or 0 hr before inoculation with CMV was suppressed. When upper leaves of tobacco plants were treated with figaren and inoculated 10 min later with CMV, mosaic symptoms were delayed for 5-7 days on most of tobacco plants, and some plants remained asymptomatic. The amount of CMV RNAs and CMV antigen in melon protoplasts inoculated with CMV and subsequently inoculated with figaren similarly increased with time as did that the control ELISA and local lesion assays indicated that CMV infection on the upper surfaces of the leaves of tobacco, melon, cowpea and Chenopodium amaranticolor whose lower surfaces had been treated with figaren 5-10 min before CMV inoculation was almost completely inhibited. Figaren almost completely inhibited CMV infection on the untreated halves of leaves of tobacco. Figaren also induced localized acquired resistance in tobacco plants. Less

克隆并测序了编码黄瓜核糖体失活蛋白（CF-RIP）的cDNA。整个cDNA含有编码286个氨基酸的蛋白质的ORF，其共享核糖体失活蛋白（RIPS）的保守残基。CF-RIP与其它四种瓜类植物RIP的同源性为54.8-64.5%。RT-PCR分析表明，CF-RIP mRNA在黄瓜叶片和茎中均有表达。经柱层析、SDS-PAGE和过碘酸-希夫染色证实，无花果素是一种分子量为23 kDa的糖蛋白。N-末端氨基酸序列分析表明，figaren含有S-等位基因相关核糖核酸酶（RNases）的保守区域。凝胶内RNase分析表明，figaren可消化酵母RNA。荧光原位杂交显示，1μ g/ml的figaren和RNase（牛胰和RNase T1）对C ...更多信息 豇豆叶片中的MV感染。无花果蛋白酶和RNases在5- 500 ng/ml浓度下对CMV的局部病斑形成有相似的抑制作用。当用无花果处理烟草植株上部叶并在10分钟后接种CMV时，大多数烟草植株的马赛克症状推迟5-7天，部分植株仍无症状。黄瓜病毒RNA和CMV抗原的量在甜瓜原生质体接种CMV和随后接种无花果类似的增加，随着时间的推移，控制ELISA和局部病变测定表明，烟草，甜瓜，豇豆和藜的叶片的上表面上的CMV感染，其下表面已被处理5-10分钟之前，CMV接种无花果几乎完全抑制。Figaren几乎完全抑制CMV感染的烟草叶片的未处理的一半。Figaren还在烟草植物中诱导局部获得抗性。少

项目成果

Fujiwara,M.: "Effect on Cucumber mosaic virus infection of Figaren, an antiviral protein from Cucumis figarei"J.Gen.Plant Pathol.. 67-2 (in press). (2001)

Fujiwara,M.：“Figaren（一种来自黄瓜的抗病毒蛋白）对黄瓜花叶病毒感染的影响”J.Gen.Plant Pathol.. 67-2（出版中）。

Saitoh,H.: "The coat protein gene is essential for the systemic infection of virus in Cucumis figarei at a high temperature"Ann Phytopath.Soc.Japan. 65・3. 248-253 (1999)

Saitoh, H.：“外壳蛋白基因对于黄瓜中病毒在高温下的全身感染至关重要”Ann Phytopath.Soc.Japan 65·3 (1999)。

Yamada,T.: "Cloning and analysis of a cDNA coding for a putative ribosome-inactivating protein from Cucumis figarei"Plant Biotechnology. 17. 337-340 (2000)

Yamada,T.：“编码来自黄瓜的假定核糖体失活蛋白的 cDNA 的克隆和分析”植物生物技术。

Murai,J.: "Isolation and characterization of the three Waxy genes encoding the granule-bound starch synthase in hexaploid wheat"Gene. 234. 71-79 (1999)

Murai，J.：“六倍体小麦中编码颗粒结合淀粉合酶的三个蜡质基因的分离和表征”基因。

Fujita,Y.: "Bromovirus movement protein gene conditions the host-specific ability of the virus to move through the vascular system and affects the ・・・"Virology. (accepted). (2000)

Fujita, Y.：“溴病毒运动蛋白基因调节病毒通过血管系统的宿主特异性能力并影响......”病毒学（已接受）。