Pyramiding of Cry Toxins and Methanol Producing Genes to Increase Insect Resistance in Cotton

The idea of enhanced methanol production from cell wall by pectin methyl esterase enzymes (PME) combined with expression of cry genes from Bacillus thuringiensis as a strategy to improve pests control in cotton is presented. We constructed a cassette containing two cry genes (cry1Fa and Cry32Aa) and two pme genes, one from Arabidopsis thaliana (AtPME), and other from Aspergillus niger (AnPME) in pCAMBIA1301 plant expression vector using CAMV-35S promoter. This construction was transformed in Eagle-2 cotton variety using shoot apex-cut Agrobacterium-mediated transformation. The expression of cry genes and pme genes was confirmed by qPCR. Methanol production was measured in control and in the cry and pme transformed plants showing methanol production only in transformed plants, then the non-transgenic cotton plants. Finally, insect bioassays performed with transgenic plants expressing cry and pme genes, showed 100 % mortality for Helicoverpa armigera (cotton bollworm) larvae, 70% mortality for pectinophore gossypiella (pink bollworm) larvae and 95% mortality of Earias fabia, (spotted bollworm) larvae, that was higher than the transgenic plants expressing only cry genes that showed 84%, 49% and 79% mortality, respectively. These results demonstrate that Bt. cry-genes coupled with pme genes is an effective strategy to improve the control of different insect pests.