Odorant receptor(OR)is crucial for insects to detect and recognize external chemical cues closely related to their survival.The insect OR forms a heteromeric complex composed of a ligand-specific receptor and a ubiqui...Odorant receptor(OR)is crucial for insects to detect and recognize external chemical cues closely related to their survival.The insect OR forms a heteromeric complex composed of a ligand-specific receptor and a ubiquitously odorant receptor coreceptor(Orco).This study used the CRISPR/Cas9 technique to knock out(KO)Orco and reveal its essential role in acting on OR-meditated olfactory behavior in a critical invasive agricultural pest,the fall armyworm(FAW),Spodoptera frugiperda.Electroantennogram(EAG)results suggested that the Orco mutants of both male and female moths severely reduced their electrophysiological responses to the eight tested plant volatiles and two sex pheromones.However,the Orco gene played distinct roles in mating behavior between sexes:the mating behavior was fully disrupted in mutated males but not in mutated females.The oviposition result indicated that the Orco KO females displayed reduced egg laying by 24.1%compared with the mated wild type(WT)females.Overall,these results strongly suggest that Orco is an excellent target for disrupting FAW’s normal behavior and provides a feasible pest control approach.展开更多
【目的】探索建立一种有效制备昆虫非典型嗅觉受体Orco抗原的方法,为Orco蛋白组织定位及功能研究奠定基础。【方法】设计带有Bam H I和Hind III酶切位点的引物,采用RT-PCR方法扩增家蚕Bombyx mori Orco第4-5跨膜区之间的基因片段,将其...【目的】探索建立一种有效制备昆虫非典型嗅觉受体Orco抗原的方法,为Orco蛋白组织定位及功能研究奠定基础。【方法】设计带有Bam H I和Hind III酶切位点的引物,采用RT-PCR方法扩增家蚕Bombyx mori Orco第4-5跨膜区之间的基因片段,将其与原核表达载体p ET-28a(+)双酶切处理后进行连接,然后转化大肠杆菌Escherichia coli感受态细胞DH5α,重组质粒再转化大肠杆菌BL21(DE3)菌株,用IPTG诱导表达,用SDS-PAGE检测诱导蛋白,用His Trap HP亲和层析对诱导表达的大量蛋白进行纯化。【结果】在大肠杆菌原核表达系统中,用IPTG诱导获得了与预测蛋白大小相符的目的蛋白,经SDS-PAGE检测发现目的蛋白以包涵体形式表达,在变性条件下经His Trap HP亲和层析获得大量可用于抗体制备的纯化蛋白。【结论】利用原核表达系统可获得制备家蚕Orco抗体的抗原蛋白。展开更多
A highly conserved and broadly expressed receptor protein Orco （olfactory coreceptor） is crucial for insect olfaction, and an orthologue of Orco has been identified in several insect species. Here we report the iden...A highly conserved and broadly expressed receptor protein Orco （olfactory coreceptor） is crucial for insect olfaction, and an orthologue of Orco has been identified in several insect species. Here we report the identification and characterization of Orco from Spodoptera litura. The protein displays high primary amino acid sequence conservation with other previously identified Orco orthologues. Bioinformatic analysis revealed that it has common features with other members of the Orco subfamily： seven-transmembrane domains with intracellular N-terminus and extracellular C-terminus. The transcript was detected in abundance in the chemosensory organs of the antennae of both male and female adults by real-time polymerase chain reaction analysis, and was localized at the bases of all categories of olfactory sensilla through in situ hybridization.展开更多
基金the National Natural Science Foundation of China(32130089)the China Postdoctoral Science Foundation(2020M680785)+3 种基金the Shenzhen Science and Technology Program,China(KQTD20180411143628272)the Central Public-interest Scientific Institution Basal Research Fund of Chinese Academy of Agricultural Sciences(CAAS-ZDRW202108)the Special Funds for Science Technology Innovation and Industrial Development of Shenzhen Dapeng New District,China(PT202101-02)the Agricultural Science and Technology Innovation Program of Chinese Academy of Agricultural Sciences(ASTIP).
文摘Odorant receptor(OR)is crucial for insects to detect and recognize external chemical cues closely related to their survival.The insect OR forms a heteromeric complex composed of a ligand-specific receptor and a ubiquitously odorant receptor coreceptor(Orco).This study used the CRISPR/Cas9 technique to knock out(KO)Orco and reveal its essential role in acting on OR-meditated olfactory behavior in a critical invasive agricultural pest,the fall armyworm(FAW),Spodoptera frugiperda.Electroantennogram(EAG)results suggested that the Orco mutants of both male and female moths severely reduced their electrophysiological responses to the eight tested plant volatiles and two sex pheromones.However,the Orco gene played distinct roles in mating behavior between sexes:the mating behavior was fully disrupted in mutated males but not in mutated females.The oviposition result indicated that the Orco KO females displayed reduced egg laying by 24.1%compared with the mated wild type(WT)females.Overall,these results strongly suggest that Orco is an excellent target for disrupting FAW’s normal behavior and provides a feasible pest control approach.
文摘A highly conserved and broadly expressed receptor protein Orco （olfactory coreceptor） is crucial for insect olfaction, and an orthologue of Orco has been identified in several insect species. Here we report the identification and characterization of Orco from Spodoptera litura. The protein displays high primary amino acid sequence conservation with other previously identified Orco orthologues. Bioinformatic analysis revealed that it has common features with other members of the Orco subfamily： seven-transmembrane domains with intracellular N-terminus and extracellular C-terminus. The transcript was detected in abundance in the chemosensory organs of the antennae of both male and female adults by real-time polymerase chain reaction analysis, and was localized at the bases of all categories of olfactory sensilla through in situ hybridization.
