Bacillus thuringiensis (Bt) toxins expressed in GM crops provide effective protection against chewing-type insects but fail to control damage by sucking pests, leading to a search for a newer class of efficient entomotoxic proteins. In this direction lectins with novel sugar-binding properties showing toxicity to a wide range of agricultural insect pests are being evaluated for the insecticidal efficacy. Our previous study demonstrated the potential toxic effect of O-glycans sugar-specific Sclerotium rolfsii lectin (SRL) supplemented through an artificial diet on the growth and survival of Spodoptera litura larvae by binding to midgut epithelial cells. Considering the significance of its entomotoxic property, we cloned the SRL gene and expressed it in a prokaryotic host, which showed 97 % sequence homology to the native SRL. Recombinant SRL (recSRL) exhibited a carbohydrate-binding property and insecticidal activity similar to native SRL. The SRL gene was transferred into tobacco plants by Agrobacterium-mediated transformation and expressed constitutively using a CaMV35S promoter. Integration and stability of the SRL gene was confirmed by Southern hybridization. recSRL expressed in the transgenic tobacco leaves was analyzed by Western blotting and quantified by ELISA, which accounted for 0.47 % of the total soluble leaf proteins. Bioassays conducted on second instar S. litura larvae fed on transgenic tobacco leaves showed 90 % larval mortality in 48 h and in planta assays using M. persicae demonstrated an 81.90 % population reduction. These findings indicated that the ectopically expressed SRL conferred protection against target pests of lepidopteran and hemipteran orders. © 2015, Springer-Verlag Berlin Heidelberg.