Identification Of Odorant Binding Proteins Genes In Apolygus Lucorum(Hemiptera: Miridae) And The Binding Specificity Analysis Of Odorant Binding Proteins AlucOBP13 And AlucOBP17

Apolygus lucorum(Meyer-Dür)(Hemiptera: Miridae) is one of the most important agricultural pests, with high mobility, broad host range and cryptic feeding habits in China. Chemosensory behavior plays an important role in many crucial stages in the life of A.lucorum, such as the detection of sex pheromone cues during mate pursuit and fragrant odorants during flowering host plant localization. OBPs are involved in the initial biochemical recognition steps in semiochemical perception. With the identification and the development of field application of sex pheromone in A. lucorum, interference of insect chemical communication has become a new pest control strategy. This research deeply exploited the mechanism of olfactory peripheral recognition between A. lucorum and sex pheromone on the molecular biology level, can contribute to lay the theoretical reference for olfactory gene RNAi or silencing and the optimization of sex pheromone components.The main contents of the experiment include two part below:1. A transcriptomics-based approach was used to identify potential OBPs in A. lucorum. In total, 38 putative OBP genes were identified, corresponding to 26 ‘classic’ OBPs and 12 ‘Plus-C’ OBPs. Phylogenetic analysis revealed that A. lucorum OBP proteins are more closely related to the OBP proteins of other mirid bugs as the same family OBP clustering together. Quantitative real-time PCR analysis for the first reported 23 AlucOBPs revealed that the expression level of 11 AlucOBP genes were significantly higher in antennae of both sexes than in other tissues. Three of them were male antennae-biased and six were female antennae-biased, suggesting their putative roles in the detection of female sex pheromones and host plant volatiles. In addition, three, four, two and one AlucOBPs had the highest degree of enrichment in the stylet, head, leg, and in abdomen tissues, respectively. Two other OBPs were ubiquitously expressed in the main tissues, including antennae, stylets, heads, legs and wings. Most orthologs had similar expression patterns, strongly indicating that these genes have the same function in olfaction and gustation.2. We screened two male-biased antennal OBPs AlucOBP13 and AlucOBP17 as the potential PBPs, which might be closely associated with the process of the detection and discrimination of sex pheromones in A. lucorum. We cloned, expressed and purified AlucOBP13 and AlucOBP17 proteins, through the fluorescence competitive binding assays and the microscale thermophoresis assays which included 9 latent sex pheromone analogs and 59 oher preferential host plant volatiles, green leaf volatiles, terpenoids and eliciting A. lucorum strong EAG response odorants. The results showed that AlucOBP13 could only bind β-ionone effectively, we considered AlucOBP13 might be a GOBP, which might have the function of locating the host plants and alteration in host ranges. AlucOBP17 not only could bind two sex pheromone analogs, hexyl hexanoate and octyl butyrate, but also bind host plant cotton volatile dodecyl aldehyde and aromatic plant volatiles β-ionone effectively, which might function in the detection and discrimination of sex pheromones, locating the host plants and alteration in host ranges.