Identification and expression analysis of a novel defense gene from Actias selene

Abstract

We isolated a novel defence gene (AsDef) cDNAs from Actias selene (Hübner 1806) (Lepidoptera: Saturniidae). It contained 691 nucleotides with a putative open reading frame of 498 bp encoding 165 amino acid residues with a reeler domain. It showed significant similarities with defence proteins from several other lepidopteran insect species including Samia ricini (Donovan 1798), Antheraea mylitta (Drury 1773), Bombyx mori (Linnaeus 1758), Lonomia oblique (Walker 1855), Hyphantria cunea (Drury 1773) and Manduca sexta (Linnaeus 1763). The recombinant protein of AsDef was expressed successfully in Escherichia coli (Migula 1895) cells, and the anti-AsDef antibody against rabbit was prepared. Results of real-time quantitative Polymerase Chain Reaction (PCR) and western blot showed that AsDef gene was detected at a particularly high levels in fat body, while AsDef protein can only be detected in haemolymph. For immune functions, AsDef gene can be upregulated by E. coli, Micrococcus luteus (Cohn 1872), Beauveria bassiana (Bals 1912) and nuclear polyhedrosis virus, and showed a stronger sensitivity for Gram-positive bacteria than fungi and virus, while the least sensitivity for Gram-negative bacteria. These data indicated that AsDef played an important role in innate immune responses of A. selene.