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棉花学报 ›› 2015, Vol. 27 ›› Issue (3): 260-267.doi: 10.11963/issn.1002-7807.201503010

• 研究与进展 • 上一篇    下一篇

大草蛉化学感受蛋白基因CpalCSP3组织表达谱及气味结合特性分析

王思豹1,2,张帅2,雒珺瑜2,李兆群1,2,吕丽敏2,王春义2,崔金杰2*,董双林1*   

  1. 1. 南京农业大学植物保护学院/农作物生物灾害综合治理教育部重点实验室,南京210095;2. 中国农业科学院棉花研究所/棉花生物学国家重点实验室,河南 安阳 455000
  • 收稿日期:2015-01-13 出版日期:2015-05-15 发布日期:2015-05-15
  • 通讯作者: cuijinjie@126.com,sldong@njau.edu.cn
  • 作者简介:王思豹(1989―),男,硕士研究生,wang_123150@163.com
  • 基金资助:
    公益性行业(农业)科研专项经费(201203036);国家现代农业产业技术体系棉花体系(CARS-18-13)

Tissue Expression Profile and Ligand Binding Affinity of the Chemosensory Protein Gene CpalCSP3 in Chrysopa pallens(Rambur)

Wang Sibao1,2, Zhang Shuai2, Luo Junyu2, Li Zhaoqun1,2, Lü Limin2, Wang Chunyi2, Cui Jinjie2*, Dong Shuanglin1*   

  1. 1. College of Plant Protection, Nanjing Agricultural University/ Key Laboratory of Integrated Management of Crop Diseases and Pests, Ministry of Education, Nanjing 210095, China; 2. State Key Laboratory of Cotton Biology / Institute of Cotton Research, Chinese Academy of Agricultural Sciences, Anyang, Henan 455000, China
  • Received:2015-01-13 Online:2015-05-15 Published:2015-05-15
  • Contact: cuijinjie@126.com,sldong@njau.edu.cn

摘要: 昆虫化学感受蛋白(Chemosensory protein, CSP)具有结合和运载气味分子穿过感器淋巴液到达嗅觉受体的作用,但尚未见有关大草蛉(Chrysopa pallens)CSP功能的报道。本研究克隆得到大草蛉CpalCSP3基因,利用RT-qPCR技术测定了其在成虫各组织的表达谱,发现CpalCSP3基因主要在雌雄虫的触角和头部表达,暗示该基因参与了大草蛉的嗅觉及味觉感受。为探讨其嗅觉功能,利用原核表达系统和荧光竞争结合试验测定了CpalCSP3蛋白与84种气味物质的结合能力。结果表明,CpalCSP3蛋白与橙花叔醇、(+) ̄雪松醇和β ̄紫罗兰酮具有很高的结合能力,结合常数(Ki)为8.79~9.79 μmol·L-1;与根皮苷、月桂醛、乙酸香叶酯和α ̄蒎烯也有较高的结合能力,Ki为11.54~19.06 μmol·L-1。分析认为,CpalCSP3蛋白可能在大草蛉对这些气味的嗅觉感受中起作用,使得大草蛉能对害虫寄主植物有效定向进而搜寻到猎物。

关键词: 大草蛉; 化学感受蛋白; 组织表达谱; 原核表达|荧光竞争结合试验

Abstract: Insect chemosensory proteins(CSPs) play important roles in olfactory perception by transporting hydrophobic chemical volatiles across the sensillum lymph to olfactory receptors, but this role in Chrysopa pallens(Rambur) has not been addressed. In this paper, a CSP gene CpalCSP3 was cloned and the tissue expression profile of the CpalCSP3 gene in male and female adults was determined by real-time quantitative polymerase chain reaction. Results showed that the CpalCSP3 gene was highly expressed in the heads and antennae of both sexes, suggesting its involvement in olfaction and gustation. To further explore the olfactory role of the gene, recombinant CpalCSP3 protein was obtained by the Escherichia coli expression system, and the binding affinities of CpalCSP3 protein for 84 plant volatiles were measured using fluorescent competitive binding assay. Among the tested compounds, nerolidol, (+)-cedrol and beta-ionone had high binding affinities to CpalCSP3 protein, with Ki values ranging from 8.79 to 9.79 μmol·L-1. In addition, phlorizin dihydrate, dodecyl aldehyde, geranyl acetate and (+/?)-alpha-pinene also showed high binding affinities, with Ki values from 11.54 to 19.06 μmol·L-1. Together, these results suggest that CpalCSP3 protein may play a role in olfaction of these plant volatiles, which facilitate host-plant searching in adult C. pallens.

Key words: Chrysopa pallens; chemosensory protein; tissue expression profile; prokaryotic expression; fluorescence competitive binding assay

中图分类号: 
  • S435.622