摘要
水稻Rho GDP解离抑制基因OsRhoGDI2是从幼穗中分离出的功能未知基因。为鉴定该基因的功能,笔者所在实验室前期构建了植物过表达载体pCAMBIA1302-OsRhoGDI2-GFP,并对水稻进行了遗传转化。对OsRhoGDI2过表达转基因水稻T_2代进行筛选和鉴定,采用PCR技术鉴定转基因植株,采用半定量RT-PCR和实时荧光定量PCR检测OsRhoGDI2在转基因水稻中的表达水平,结果显示,其中6个株系为过表达转基因植株,OsRhoGDI2表达水平上调1.69~13.35倍。为检测外源基因在转基因水稻中的拷贝数,分别以蔗糖磷酸合成酶基因SPS和潮霉素抗性基因HYG为内参基因和标记基因,采用实时荧光定量PCR(qPCR)技术结合内参基因和标记基因的标准曲线进行分析,结果显示在所检测的6个转基因株系中,外源基因的拷贝数均为1,提示已经获得稳定遗传的OsRhoGDI2过表达转基因水稻,为后续OsRhoGDI2基因的功能研究奠定基础。
引文
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