›› 2017, Vol. 60 ›› Issue (12): 1403-1410.doi: 10.16380/j.kcxb.2017.12.006

• RESEARCH PAPERS • Previous Articles     Next Articles

Cloning and mRNA expression analysis of RBP1 gene in Apis mellifera (Hymenoptera: Apidae)

HU Wan-Wan1,#, LI Shu-Yun1,2,#, ZENG Zhi-Jiang1, YAN Wei-Yu1, WANG Zi-Long1,*    

  1.  (1. Honeybee Research Institute, Jiangxi Agricultural University, Nanchang 330045, China; 2. Ganzhou Agricultural College, Ganzhou, Jiangxi 341100, China)
  • Online:2017-12-20 Published:2017-12-20

Abstract: 【Aim】 RBP1 is an important splicing factor involved in alternative splicing of the pre-mRNA of Drosophila melanogaster sex-determining gene dsx. This study aims to clone the nucleotide sequence of RBP1 gene AmRBP1 of Apis mellifera, and to analyze the domains of its amino acid sequence and its mRNA expression profiles in different developmental stages of embryo and larva, so as to provide a basis for studying its function in honeybee sex determination. 【Methods】 Several transcriptome sequences of AmRBP1 were obtained by homology search with Drosophila melanogaster rbp1 based on the predicted gene database and the transcriptome database of A. mellifera, and PCR primers were designed based on these transcriptome sequences. Then, AmRBP1 was cloned and its nucleotide and amino acid sequences were analyzed with bioinformatic software. The expression profiles of AmRBP1 in different developmental stages of embryo and larva of A. mellifera were assayed by semi-quantitative RT-PCR. 【Results】 The results showed that AmRBP1 has three alternative splicing variants named AmRBP1-R1 (GenBank accession number:KY404154), AmRBP1-R2 (GenBank accession number: KY404155) and AmRBP1-R3 (GenBank accession number: KY404156), respectively, whose mRNA sequences are 696, 790 and 771 nt in length, encoding proteins of 130, 166 and 162 aa, respectively. Protein domain prediction indicated that all of the three isoforms contain the same domains, with an amino terminal RRM domain and a carboxyl terminal RS domain. Homology analysis showed that AmRBP1 has 96.97%, 94.20%, 84.85%, 81.48%, 80.28%, 78.15% and 69.23% amino acid sequence identity with RBP1 proteins of Nasonia vitripennis, Solenopsis invicta, Aedes aegypti, D. melanogaster, Bombyx mori, Musca domestica and Anopheles sinensis, respectively. The semi-quantitative RT-PCR results showed that AmRBP1-R1-3 were expressed in various developmental stages of embryo and larva, with no obvious sex difference. 【Conclusion】AmRBP1 may be a member of SR family splicing factors with no obvious sex difference, and whether it is involved in the sex-specific splicing of doublesex (dsx) pre-mRNA needs further research.

Key words: Apis mellifera, gene cloning, expression analysis, sex determination, alternative splicing