美洲商陆抗真菌蛋白转化烟草的研究和抗病性检测

本研究为美洲商陆抗病毒蛋白(PaAFP)基因首次对植物遗传转化的研究,转入烟草中研究此蛋白对烟草立枯病的抗性。从美洲商陆叶片中获得美洲商陆抗真菌蛋白前体蛋白基因cDNA序列,构建植物表达载体pCAMBIA1300-PaAFP,通过三亲杂交法将其导入根癌农杆菌LBA4404受体菌,转染烟草获得了大量再生转基因植株。PCR、Southern杂交、RT-PCR以及Tris-Tricine-SDS-PAGE检测结果表明目的基因已经整合到烟草基因组中,并且已经得到转译。转基因植株苗期抗立枯病试验表明,转基因烟草植株对立枯丝核菌表现出了抗性。

Study on Transformation of PaAFP Gene into Tobacco

Plant expression vector pCAMBIA1300-PaAFP was constructed by cloning PaAFP gene of P.americana to pCAMBIA1300,and then the engineered strain LBA4404 is constructed by transforming plant expression vector pCAMBIA1300-PaAFP into A.tumefaciens LBA4404,which was used to transform tobacco.PCR,Southern blotting,RT-PCR and Tris-Tricine-SDS-PAGE analysis indicated that PaAFP gene had been integrated in tobacco genome and translated.Infection testing indicated that parts of PaAFP transgenic tobacco plants appeared resistant to Rhizoctonia solani.These transgenic tobacco plants had many white spots on their aged leaves.