Abstract: 【Objective】The objective of this study is to clone the cDNA sequence of AcerOBP14 from Apis cerana cerana, predict the protein structure, explore its expression profiles in different tissues and at different developmental stages of A. c. cerana, and to provide a fundamental evidence for the future study of the physiological function of this gene.【Method】 The full-length cDNA sequence of AcerOBP14 was cloned from the entire tissues of A. c. cerana by RT-PCR and submitted to the GenBank database. Physiochemical properties and structure characteristics of the deduced amino acid sequence were analyzed by multiple bioinformatics methods. Phylogenetic tree between AcerOBP14 and its homologous OBPs from Hymenoptera insects was constructed using neighbor-joining of MEGA5.2. The expression levels of AcerOBP14 mRNA in different tissues (antenna, head, thorax, abdomen, legs and wings) at 1, 5, 10, 15, 20, 25 and 30 days were detected by real-time PCR.【Result】The full-length cDNA sequence of AcerOBP14 was obtained (GenBank accession number is KT24648). The open reading frame (ORF) of AcerOBP14 is 408 bp, encoding a protein of 135 aa with an estimated molecular weight of 15.08 kD and pI of 5.44. The encoding protein has a signal peptide and no transmembrance structure, possesses a PBP-GOBP superfamily domain between 18-127 residues, exists multiple obviously hydrophobic regions binding liposoluble odorant and contains four conserved cysteins, suggesting that it belongs to the Minus-C OBP subfamily. Subcellular localization results showed that AcerOBP14 was located on the secretory pathway about endoplasmic reticulum, golgi and plasmalemma which belongs to the secretory protein. AcerOBP14 possesses 7 α-helixs, among which the α1-α6 belong to the core helixs of classical OBPs and α7 is exposed to the surface of protein at the C-terminus. The results of amino acid sequence alignment showed that AcerOBP14 was homologous to AmelOBP14 with 86% amino acid sequence identity, and then had 55% sequence identity with AdorGOBP56a-like. Phylogenetic analysis revealed that AcerOBP14, AmelOBP14, AdorGOBP56a-like, AcerOBP21 and AfloGOBP56d-like belonging to the Apis were clustered into one group, MrotGOBP56a-like (Meqachile), MdemGOBP83a-like (Microplitis), OcorOBP3 (Osmia), BterGOBP56d-like (Bombus) and BimpGOBP56d-like (Bombus) were clustered into the other group. Besides, the results of real-time PCR showed that the AcerOBP14 transcripts were differentially expressed in the antenna, head, thorax, abdomen, legs and wings of the adult workers. The thorax of 1-day-old workers had the highest expression level of AcerOBP14, the antenna had the second highest expression level, and both of them were significantly higher than the head, abdomen, legs and wings (P＜0.01). Moreover, the expression profiles in worker bees at other days were significantly higher in antenna than that in other tissues (P＜0.01), of which the antenna of 20-day-old workers had the highest expression level. According to the expression profiles of AcerOBP14 at different developmental stages, the antenna of workers had the lowest expression level at 1-day-old which was significantly lower than other days (P＜0.01), increased gradually at 5-day-old and 10-day-old, then decreased suddenly at 15-day-old. The expression level at 20-day-old was significantly higher than other days (P＜0.01), afterwards, the expression level gradually reduced. Generally, the expression profiles of the head, thorax, abdomen, legs and wings appeared a downward trend with the days of age increasing, and maintained a lower level after decreasing remarkably at 5-day-old.【Conclusion】AcerOBP14 belongs to the Minus-C OBP subfamily with a PBP-GOBP superfamily domain. The results of expression profiling suggested that AcerOBP14 belongs to general odorant binding protein (GOBP) and is involved in not only the olfactory recognition, but also other physiological functions.

Key words: Apis cerana cerana, AcerOBP14, gene cloning, bioinformatics analysis, temporal-spacial expression