Thesis (M.Sc.) - Cairo University - Faculty of Agriculture - Department of Genetics

Vacuolar - H⁺ ATPase (V - ATPase) is a multi subunit enzyme. It consists of two domains (V₁ and V₀). V₁ domain has eight subunits (A - H) while VO domain has six subunits (a, c, c', c'', d and e). The V - ATPase is an ATP - driven proton pump and is essential to the function of eukaryotic cells. In insects, V - ATPase is located in apical membrane of goblet cells in the midgut. It acts as an energizer of the plasma membrane, driving nutrient uptake, fluid secretion and in some cases alkalizing the gut lumen. The aim of the present study was knocking down the V - ATPase gene(s) using RNAi in an attempt to disturb the food process within the pink bollworm (PBW) midgut and eventually causing the insect death. Based on the conserved regions of the V - ATPase subunits A and D genes in different insects available in the Gen Bank, degenerate primers were synthesized and used to amplify the two genes. The amplified genes encoding the V - ATPase subunits A and D transcripts were sequenced. The sequencing results confirmed the isolation of the full length genes encoding the V - ATPase subunit A was 2602 bp in length, encoding 618 amino acids. While, the gene encoding V - ATPase subunit D was 1467 bp, encoding 249 amino acids. Three dsRNA fragments were designed, two of them (VATPA756 - 1155 bp and VATPA347-753 bp) were targeting subunit A and the third - VATPD221 - 703 bp to knockdown subunit D

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