甜菜夜蛾触角气味受体基因OR18的克隆和表达定位

Cloning and Localization of an Odorant Receptor Gene OR18 in the Antenna of Spodoptera exigua

【目的】从甜菜夜蛾(Spodoptera exigua)成虫触角中克隆气味受体基因,研究受体基因在虫体不同组织和触角不同感受器中的表达分布,从而探讨受体基因的功能。【方法】通过PCR结合RACE技术克隆气味受体基因的全长序列,利用实时定量PCR检测其在不同组织中的表达,采用原位杂交确定其在触角不同感受器中的分布。【结果】通过同源克隆的方法从甜菜夜蛾触角中获得1条740 bp的基因片段,通过RACE技术获得全长序列并命名为SexiOR18(GenBank登录号JN873314)。SexiOR18 cDNA全长1 618 bp,开放阅读框长度为1 194 bp,编码398个氨基酸。序列比对和进化树分析结果显示SexiOR18与其它鳞翅目昆虫尤其是夜蛾科昆虫的OR18具有很高的相似性。实时定量PCR检测结果显示,SexiOR18主要在成虫触角中表达,且在雌虫中的表达量显著高于雄虫,SexiOR18在成虫其它组织和幼虫触角中无明显表达。原位杂交结果显示,SexiOR18主要在毛形感器和锥形感器下表达,而在腔锥形感器和刺形感器下没有表达。【结论】SexiOR18在感受性信息素和普通气味的毛型感器和锥形感器中都有分布,推测其可能参与了性信息素和普通气味分子的识别。

[Objective] The objective of this study is to clone odorant receptor gene and investigate its expression in different tissues and antennal sensilla of Spodoptera exigua adults, and thus to explore the function of the odorant receptor gene. [Method] The full length cDNA sequence encoding SexiOR18 was cloned by homologous-based PCR and RACE techniques. The expression level of this gene in different tissues was investigated by quantitative real-time PCR. The in situ hybridization was used to examine its localization in different antennal sensilla of adult antennae. [ Result] A 740 bp fragment of a OR candidate gene was obtained from the antenna of S. exigua by using homologous-based cloning strategy. The full-length cDNA sequence encoding this gene was further obtained by using RACE technique. The gene was named as SexiOR18 （GenBank accession number： JN873314）. The full-length cDNA of SexiOR18 is 1 618 bp with an open reading frame of 1 194 bp, which encodes a polypeptide of 398 amino acids. The multiple alignment and phylogenetic tree analyses showed that SexiOR18 shared high identity with the homologs from other Lepidopteran species, especially noctuid insects. The expression measurements revealed that SexiOR18 expressed in significantly higher level in female antenna than in male antenna, and within the antenna, the expression was mainly restricted under trichoidsensilla and basiconic sensilla, but no expression under coeloconic sensilla and chaetica sensilla. [Conclusion] The SexiOR18 localizes under both trichoid sensilla and basiconic sensilla, suggesting its involvement in detection of both pheromone and general odorants.