Expression and inheritance of chimeric cry2AX1 gene in transgenic cotton plants generated through somatic embryogenesis

Abstract

Transgenic cotton plants expressing a novel cry2AX1 gene consisting of sequences from the cry2Aa and cry2Ac genes, driven by the CaMV35S or the EnCaMV35S promoters, were generated through Agrobacterium-mediated transformation. Among 40 independent PCR-positive events, 14 produced Cry2AX1 protein at detectable levels. Cry2AX1 levels in T0 plants ranged from 0.012 to 1.068 μg/g fresh leaf tissue. Insect bioassays on Cry2AX1-expressing T0 transgenic lines showed 27.0 to 88.8% mortality of Helicoverpa armigera. Southern hybridization analysis revealed single locus integration of the cry2AX1 gene in two T0 plants which caused 87.5 and 88.8% mortality in H. armigera. The presence and expression of the cry2AX1 gene were confirmed in T1 progeny using PCR and quantitative ELISA. The highest expression level of Cry2AX1 protein recorded in the T1 generation was 0.766 μg/g fresh leaf tissue, far less than the expression of Cry2Ab protein (23 μg/g leaf tissue) reported in the commercialized Bt cotton, Bollgard II. In spite of the low Cry2AX1 level, the mortality against H. armigera ranged between 33.3 and 85.7% in the T1 generation. These results indicate that this novel cry2AX1 gene produced a potent insecticidal protein that was effective against H. armigera in transgenic cotton plants at very low concentrations.