Cloning and expression analysis of the foraging gene in the Chinese honeybee (Apis cerana cerana Fabricius)
Author of the article:MA Wei-Hua1, 2** SHAO You-Quan1 ZHAO Hui-Ting3 MENG Jiao2 TIAN Song-Hao4 DU Ya-Li2 JIANG Yu-S
Author's Workplace：1. Institute of Horticulture, Shanxi Academy of Agricultural Science, Taiyuan 030031, China; 2. College of Animal Science and Veterinary Medicine, Shanxi Agricultural University, Taigu 030801, China; 3. College of Life Science, Shanxi Agricultural University, Taigu 030801, China; 4. Fenyang College of Shanxi Medical University, Fenyang 032200, China
Key Words：Chinese honeybee, foraging gene, cloning, development expression pattern, PKG activity
[Objectives] To lay a foundation for studying the molecular mechanisms by which bee genes regulate the division of labor and foraging behavior. [Methods] The cDNA sequence of Acfor from the brain of Apis cerana cerana Fabricius was cloned using the Reverse transcription-polymerase chain reaction (RT-PCR) and its protein structure analyzed using bioinformatics software. A quantitative analysis of its expression in worker bees on successive days was conducted by the quantitative real-time PCR (qRT–PCR). The activity of cGMP-dependent protein kinase (PKG) was quantified with the colorimetric method. [Results] The results show that the full-length cDNA sequence of Acfor is 3 029 bp (GeneBank accession no.KP662686.1), including an open reading frame (ORF) of 2 169 bp that encodes 722 amino acids. The molecular weight and predicted isoelectric point of this protein are 81.80 ku and 5.50, respectively. The protein is water-soluble, acidic and stable, has no signal peptide or transmembrane domain, and has 2 glycosylation sites and 39 phosphorylation sites that may be located in the cytoplasm. The predicted secondary structure shows that Acfor protein is comprised of 57.06% random coil, 28.12% alpha helix and 14.82% extended strand. Phylogenetic analysis indicates that Acfor of A.c.cerana is in the same clade as similar proteins of other Hymenopteran insects. This clade has seven sub-branches, and the protein with the closest molecular distance to Acfor is Amfor. Acfor transcript expression and PKG activity were detected in the heads of workers of different ages. Temporal trends of Acfor mRNA expression and PKG activity were similar. Gene expression and PKG activity decreased from day 1 to 10, and begin to rise after day 10, remaining at peak levels until day 25 after which they declined with increasing age. [Conclusion] Acfor expression and PKG activity levels affect A. c. cerana age-related foraging behavior.