Tiwari Siddharth, Mishra Devesh K, Singh Ankit, Singh P K, Tuli Rakesh
Plant Molecular Biology and Genetic Engineering Division, National Botanical Research Institute, Rana Pratap Marg, Lucknow, UP 226001, India.
Plant Cell Rep. 2008 Jun;27(6):1017-25. doi: 10.1007/s00299-008-0525-x. Epub 2008 Mar 5.
The tobacco cutworm (Spodoptera litura) is a polyphagous foliage insect and a major pest on peanut (Arachis hypogaea L.). S. litura is susceptible to the chimeric delta-endotoxin Cry1EC reported earlier. De-embryonated cotyledon explants of peanut were transformed using Agrobacterium tumefaciens strain EHA101 harboring a synthetic cry1EC gene driven by the CaMV 35S promoter. Transgenic plants of peanut with a single copy insertion of cry1EC were selected in the T(0) generation by Southern blot hybridization. Real-time PCR, Western blot and ELISA analysis indicated that expression of the cry1EC gene was higher in single copy T(1) plants. Immunoassay showed expression of Cry1EC up to 0.13% of total soluble protein in T(1) plants. Leaf feeding bioassay on highly expressing transgenic lines showed 100% killing of larvae at the 2(nd) instar stage of S. litura. This is the first report of transgenic peanut plants with resistance to S. litura.
烟草夜蛾（斜纹夜蛾）是一种多食性食叶昆虫，也是花生（落花生）的主要害虫。斜纹夜蛾对先前报道的嵌合δ-内毒素Cry1EC敏感。使用携带由花椰菜花叶病毒35S启动子驱动的合成cry1EC基因的根癌农杆菌EHA101菌株转化花生的去胚子叶外植体。通过Southern印迹杂交在T(0)代中筛选出cry1EC单拷贝插入的花生转基因植株。实时PCR、Western印迹和ELISA分析表明，cry1EC基因在单拷贝T(1)植株中的表达较高。免疫测定显示，T(1)植株中Cry1EC的表达量高达总可溶性蛋白的0.13%。对高表达转基因株系进行的叶片喂食生物测定表明，在斜纹夜蛾幼虫的二龄阶段，幼虫死亡率达100%。这是关于抗斜纹夜蛾花生转基因植株的首次报道。
