The successful deployment of insecticidal proteins from Bacillus thuringiensis (Bt) via genetically engineered crops has revolutionized pest management practices in agriculture. However, the development of insect resistance to Bt toxins threatens the sustainable application of Bt-biotechnology. Current understanding of Bt resistance has indicated that resistance to Bt in insects is complex, involving multiple midgut proteins that serve as receptors for Bt toxins in the intoxication pathways. In this dissertation, I studied the genetic basis of resistance to Bt protein Cry1Ac in a greenhouse-evolved resistant strain of the cabbage looper, Trichoplusia ni. Using genetic and molecular approaches, I studied the genetic association of altered APN (Aminopeptidase N) expression with the ABCC2 (ABC transporter C2) and the association of Cry1Ac resistance with APNs. The results indicated that the downregulated APN1 expression in resistant T. ni and low expression of APN6 in susceptible T. ni were associated with mutations in APN1 and APN6 genes but were not with ABCC2. The high-level resistance to Cry1Ac in T. ni is associated with both ABCC2 and APN1 mutations and additional factors to be identified. The functional roles of putative Cry1Ac receptors ABCC2, ALP (alkaline phosphatase), APN1, and midgut cadherin (CAD) in larval susceptibility to Cry1Ac were systematically examined, using a series of gene knockout T. ni mutant strains. ABCC2 was confirmed to be a major receptor for Cry1Ac in T. ni, but knockout mutations in the ALP, APN, and CAD receptors resulted in none to low resistance in T. ni. Results also indicated additional resistance genes to be identified. Finally, I studied the role of carbohydrate moieties in the mode of action of Cry1Ac. The interaction of Cry1A with the midgut and Cry1Ac toxicity in larvae were analyzed in vitro and in vivo, using a carbohydrate binding chemical, Calcofluor. The results demonstrated that carbohydrate moieties play critically important roles in the functional specific binding of the toxin to the midgut receptors in the pathway of toxicity. Overall, the findings from this thesis research advanced our understanding of the mode of action of Cry proteins and mechanisms of insect resistance to Cry proteins.