【目的】从绿盲蝽Apolygus lucorum触角中克隆气味受体基因Aluc OR40,研究该气味受体基因在绿盲蝽不同组织中的表达水平,然后对该基因的功能进行研究,为理解绿盲蝽的嗅觉识别机制提供理论基础。【方法】在绿盲蝽成虫触角转录组测序与分析的基础上,通过PCR技术克隆得到气味受体基因Aluc OR40的全长序列。用半定量RT-PCR研究该基因在雌雄虫不同组织中的表达水平。然后通过爪蟾卵母细胞体外表达该基因,并结合双电极电压钳检测了该气味受体对60种气味分子包括绿盲蝽性信息素组分和植物挥发物的反应。然后,利用触角电位技术测定羽化后3 d的绿盲蝽成虫对Aluc OR40的气味配体的触角电位反应。【结果】克隆了Aluc OR40(Gen Bank登录号:KU886190)。Aluc OR40编码398个氨基酸,蛋白具有7个跨膜结构域,N末端位于胞内,C端位于细胞外,符合昆虫气味受体的典型特征。半定量RT-PCR的结果显示,Aluc OR40在触角中特异表达,且在雄虫触角中的表达水平高于雌虫。双电极电压钳记录结果显示,Aluc OR40只对反-2-己烯醇一种气味分子具有特异反应。EAG实验结果表明,羽化后3 d的绿盲蝽雌雄虫都对反-2-己烯醇有反应,且雄虫触角的EAG反应显著高于雌虫。【结论】结果提示Aluc OR40参与了绿盲蝽对反-2-己烯醇的识别过程,推测其在绿盲蝽交配过程中雄虫对雌虫的识别中发挥作用。
【Aim】This study aims to clone an odorant receptor gene,Aluc OR40,from Apolygus lucorum,to characterize its expression profiles in different tissues and to explore its function,so as to provide a theoretical basis for understanding the mechanisms of odor reception in A. lucorum. 【Methods 】The antennal transcriptome sequencing and RT-PCR amplification was used to clone the full-length c DNA of Aluc OR40. The relative expression levels of this gene in different tissues of male and female adults of A.lucorum were detected using semi-quantitative RT-PCR. Through the combination of heterologous expression in Xenopus oocytes and two-electrode voltage clamp electrophysiological recording technique,the response of this odorant receptor to 60 odorants including sex pheromone of A. lucorum and plant volatiles was determined. The EAG responses of the 3-d-old adults of A. lucorum to the odorant ligand of Aluc OR40 were further measured. 【Results】Aluc OR40( Gen Bank accession number: KU886190) was cloned. Aluc OR40 encodes 398 amino acids and the deduced protein has 7 transmembrane domains with an intracellular N-terminus and an extracellular C-terminus,which conforms to the typical structure of insect odorant receptors. The results of semi-quantitative RT-PCR showed that Aluc OR40 was specifically expressed in the antenna of adults and the expression level was obviously higher in males than females.Two-electrode voltage clamp electrophysiological recording showed that Aluc OR40 specifically responded to( E)-2-hexenol among the tested odorants. The EAG experiments showed that the 3-d-old adults had EAG response to( E)-2-hexenol,but the responses of males were significantly higher than those of females. 【Conclusion】Our results suggest that the Aluc OR40 is involved in identification of( E)-2-hexenol and may play an important role in identification of female by male during mating process.