Development of broad-spectrum and sustainable resistance in cotton against major insects through the combination of Bt and plant lectin genes.

KEY MESSAGE: Second generation Bt insecticidal toxin in comibination with Allium sativum leaf agglutinin gene has been successfully expressed in cotton to develop sustainable resistance against major chewing and sucking insects. The first evidence of using the Second-generation Bt gene in combination with Allium sativum plant lectin to develop sustainable resistance against chewing and sucking insects has been successfully addressed in the current study. Excessive use of Bt δ-endotoxins in the field is delimiting its insecticidal potential. Second-generation Bt Vip3Aa could be the possible alternative because it does not share midgut receptor sites with any known cry proteins. Insecticidal potential of plant lectins against whitefly remains to be evaluated. In this study, codon-optimized synthetic Bt Vip3Aa gene under CaMV35S promoter and Allium sativum leaf agglutinin gene under phloem-specific promoter were transformed in a local cotton variety. Initial screening of putative transgenic cotton plants was done through amplification, histochemical staining and immunostrip assay. The mRNA expression of Vip3Aa gene was increased to be ninefold in transgenic cotton line L6P3 than non-transgenic control while ASAL expression was found to be fivefold higher in transgenic line L34P2 as compared to non-transgenic control. The maximum Vip3Aa concentration was observed in transgenic line L6P3. Two copy numbers in homozygous form at chromosome number 9 and one copy number in hemizygous form at chromosome number 10 was observed in transgenic line L6P3 through fluorescent in situ hybridization. Significant variation was observed in transgenic cotton lines for morphological characteristics, whereas physiological parameters of plants and fiber characteristics (as assessed by scanning electron microscopic) remained comparable in transgenic and non-transgenic cotton lines. Leaf-detach bioassay showed that all the transgenic lines were significantly resistant to Helicoverpa armigera showing mortality rates between 78% and 100%. Similarly, up to 95% mortality of whiteflies was observed in transgenic cotton lines when compared with non-transgenic control lines.