冈田绕眼果蝇PoOBP56基因的鉴定、生物信息学及组织表达模式分析

Identification,bioinformatic analysis,and analysis of the pattern of tissue expression of the PoOBP56 gene in Phortica okadai

目的鉴定冈田绕眼果蝇(Phortica okadai)PoOBP56基因,并对其进行生物信息学及组织表达模式分析。方法从P.okadai头部转录组数据中筛选出高表达量基因PoOBP56,并对其进行PCR验证;基于生物信息学方法分析PoOBP56编码蛋白的结构特征,包括PoOBP56蛋白基本理化性质、蛋白疏水性、蛋白跨膜区域、蛋白信号肽结构、蛋白功能域、蛋白糖基化位点、磷酸化位点及其二、三级结构,并构建系统进化树;应用RT-qPCR(实时荧光定量PCR)检测该基因在蝇体组织部位的表达差异。结果从冈田绕眼果蝇头部转录组中筛选出1条具有完整编码区的气味结合蛋白(Odorant-binding protein,OBPs)基因序列,该基因包含806 bp的开放阅读框,编码143个氨基酸,蛋白相对分子质量为66.5×10^(3),无跨膜结构。同源序列比对显示,该蛋白氨基酸序列与斑翅果蝇(Drosophila suzukii)、Drosophila biarmipes等科果蝇相似性高,与黑腹果蝇(Drosophila melanogaster)相似性存在差异。RT-qPCR检测雌蝇PoOBP56基因触角表达量较高,雄蝇触角、蝇足和蝇翅的表达量较高,雄翅表达量高于雌翅(P<0.05),雌雄其余相同部位间表达量差异无统计学意义(P>0.05)。结论本研究筛选出冈田绕眼果蝇PoOBP56基因,该基因主要在其触角高表达,编码蛋白与斑翅果蝇高度同源,为进一步揭示果蝇-环境间的化学通讯机制以及蝇媒防治措施的制定奠定了基础。

Objective To identify the PoOBP56 gene of Phortica okadai and analyze its bioinformatics and pattern of tissue expression.Methods The highly expressed gene PoOBP56 was screened from data on the P.okadai head transcriptome,and it was verified using PCR.The structural characteristics of the PoOBP56 encoded protein were bioinformatically analyzed,including the basic physical and chemical properties of the protein,its hydrophobicity,transmembrane regions,signal peptide structure,functional domains,glycosylation sites,phosphorylation sites,and secondary and tertiary structure.In addition,a phylogenetic tree was also constructed.Real-time fluorescent quantitative PCR(RT-qPCR)was used to detect the differential expression of the PoOBP56 gene in various tissues of the fly body.Results An OBP gene with a complete coding region was screened from the head transcriptome of P.okadai.This OBP gene contained an 806-bp open reading frame,it encoded 143 amino acids,and it had a molecular weight of 66.5×10^(3) without a transmembrane structure.The amino acid sequence of P.okadai was highly similar to that of Drosophila suzukii and D.biarmipes but differed from that of D.melanogaster.RT-qPCR indicated that levels of expression of the PoOBP56 gene in females were higher in the antennae while levels of expression in the antennae,legs,and wings were higher in males.Levels of expression were higher in the wings of males than in the wings of females(P<0.05).There were no significant differences in levels of expression in other parts of males and females(P>0.05).Conclusion The PoOBP56 protein was screened from P.okadai.The gene was highly expressed in the antennae,and the coding protein was highly homologous to D.suzukii.These findings have laid the foundation to further reveal the mechanism of chemical communication between Drosophila and the environment and to formulate measures to control Drosophila as a vector.