关键词: 粘虫, 类钙粘蛋白基因, 基因克隆, 序列分析, 蛋白结构, 时空表达

Abstract:  The cadherin-like proteins, located in the brush border membrane vesicles (BBMV) of insect midgut, are major receptors of Bacillus thuringiensis  insecticidal crystal protein. The complete cDNA sequence of cadherin-like protein gene was amplified from the oriental armyworm, Mythimna separata, using RT-PCR and RACE techniques. The full cDNA sequence (named as Msclp, GenBank accession no. JF951432) is 5 642 bp in length, encoding 1 757 amino acid residues, and the deduced amino acid sequence shows typical characteristics of the known insect cadherin proteins, including a signal peptide, a proprotein region, twelve cadherin repeats, a membrane-proximal region, a transmembrane region and a cytoplasmic region. The predicted molecular mass and isoelectric point are 196.786 kD and 4.5, respectively. The cadherin-like protein from M. separata shows the close relationship to cadherin proteins from Helicoverpa assulta, Heliothis virescens, Helicoverpa armigera and Spodoptera exigua, with the amino acid identities of 61.77%, 61.66%, 61.26% and 58.14%, respectively. The real-time RT-PCR results showed that relative expression levels of cadherin-like gene were significantly different in different instars of larvae (P<0.01), which was the highest in the 4th instar larvae and the lowest in the newly-hatched larvae. The cadherin-like gene was majorly expressed in the midgut and very lowly expressed in other parts of the larval body. The results provide the basis for revealing the insecticidal mechanism of Bt to oriental armyworm, and the resistance mechanism of oriental armyworm to Bt.

Key words: Mythimna separata, cadherin-like gene, gene cloning, sequence analysis, protein structure, spatio-temporal expression