Abstract: 【Aim】 The objective of this study is to analyze the distribution of AhetPBP2 in the antenna of Atrijuglans hetaohei. 【Methods】 The cDNA was synthesized with the total RNA extracted from antennae of newly emerged adults of A. hetaohei. The cDNA segment of AhetPBP2 was amplified by RT-PCR with degenerate primers. The full-length cDNA sequence of AhetPBP2 was obtained by RACE. AhetPBP2 without the signal peptide sequence was expressed in Escherichia coli BL21(DE3). Recombinant AhetPBP2 was purified through Ni-chelating affinity chromatography. The polyclonal antiserum was prepared by immunizing a male New Zealand white rabbit with the purified AhetPBP2. Immunohistochemical analysis was performed with the polyclonal antibody of AhetPBP2. 【Results】 The full-length cDNA sequence of AhetPBP2 is 923 bp. Its open reading frame is 504 bp, encoding a protein of 167 amino acids with the molecular weight of 19.26 kD and the isoelectric point of 5.47. After induction with 1 mmol/L IPTG for 10 h, the soluble recombinant protein AhetPBP2 was obtained. Western blot showed that the recombinant protein was expressed successfully. ELISA detection showed that the titer for the polyclonal antibody was 1:1 024 000. Immunohistochemical analysis showed that parts of the antennal sensilla were labeled by AhetPBP2 polyclonal antibody. 【Conclusion】 AhetPBP2 is localized on parts of antennal sensilla of A. hetaohei adults, suggesting that these antennal sensilla may be involved in the perception of sex pheromones.

Key words: Atrijuglans hetaohei, sex pheromone binding protein, prokaryotic expression, polyclonal antibody, immunofluorescence localization