Inheritance Of Resistance To Cry1Ab Toxin In Ostrinia Furnacalis And Cloning The Cadherin-like Gene In Midgut

Asian corn borer, Ostrinia furnacalis (Guenee), (ACB), is the most important insect pest on maize in China. Athough transgenic Bt maize that expresses CrylAb toxin can provide season long protectionfrom this insect, its success will be short-lived if ACB adapt to Bt maize. A resistant strain to Cryl Ab ofACB has been developed through laboratory screening. In order to develop a functional resistantmanagement strategy, it is important to understand the pattern of inheritance of resistance. In addition,understanding the molecular mechanism of resistance to CrylAb in ACB is a basis of resistancemanagement, especially for developing a practical method in detecting and monitoring the evolution ofACB resistance to Bt maize in the field.The mode of inheritance to CrylAb toxin was elucidated through bioassay analysis of the responseof resistant, susceptible, reciprocal F₁ hybrids and backcross [BC₁(♀_SR×♂_SS) and BC₂(♀_RS×♂_ss)] ACBprogenys to CrylAb using semi-artifical diet. The median lethal concentrations (LC₅₀s) for susceptible,resistant, F1, and backcross progenys were 2.54, 45.45, 14.86(SR), 19.57 (RS), 9.68 (BC₁), and9.60μg/g (BC₂), and IC50_s 0.29, 5.32, 2.14, 2.37, 1.18, and 0.91μg/g. The progeny of reciprocal F₁hybrids responded alike in bioassays, indicating autosomal inheritance. The degree of dominance valuesfor SR and RS were 0.22 and 0.41 based on LC₅₀, and 0.37 and 0.43 based on IC₅₀, the estimates ofdominance were 0.61 and 0.70, 0.68 and 0.71, respectively. The values of x² for BC₁ and BC₂ were 8.68and 8.36, lower than x²_0.05. It suggests that resistance to CrylAb was autosomal and inherited as anincomplete dominant trait, governed by single gene.By designing the degenerate primers and using RT-PCR and RACE methods, the cDNA geneencoding cadherin-like protein of ACB was cloned and sequenced. It was 5570 bp, 5'and 3'UTR were222bp and 196bp, a single 5154bp open reading frame encoded 1717 amino acid polypeptide, molecularweight was estimated as 192kDa, predicted isoelectric point was 4.21. The amino acid sequence wassimilar to O. nubilalis, involving a 21 residue single peptide (SIG), eleven consensus cadherin repeats(CR), membrane proximal region (MPR), 23 residue transmembrane region (TMR) and a smallcytoplasmic region (CPR). There were 14 predicted cadherin peptide N-glycosylation sites. Thehomologous between O. furnacalis and O. nubilalis was 99.0%, and more than 50% with otherlepidopteran insects.In comparison with the cadherin-like gene sequence in susceptible strain, 40 nucleotide mutationswere observed in resistant strain, which contriduted to 13 amino acid differences (K³⁰⁴→E, C⁴²⁷→Y,V⁵⁴²â†’E, R¹⁰⁰³â†’Q, Q¹⁰⁹³â†’p, H¹²⁵³â†’R,D¹²⁷¹â†’Y, V¹³³⁸→I, D¹³⁷⁹→N, I¹³⁸⁴→T, T¹⁴⁵⁷→S, V¹⁶⁸³â†’I,S¹⁶⁹⁷→T) in the putative protein sequences. The characters of 8 amino acids were changed, 6 of themwere located in 1200-1470 polypeptide region. These mutations may associate with the resistance ofACB to Bt CrylAb.