Identification Of Oflactory Related Genes And Functional Analysis Of Odorant Binding Proteins In Apriona Germari(Hope)

Apriona germari(Hope)is a destructive wood-boring pest,and their lives are hidden and difficult to prevent and control.Traditional control methods can not meet the existing control needs,and chemical ecological control technology has become an important way to control longicorn beetles.Studying the olfactory response mechanism of A.germari has important theoretical significance for the development of green prevention and control technologies.When responding to semiochemicals,the longicorn beetle utilizes not only its main sensory organ,the antennae,but also its maxillary palps,labial palps,and reproductive organs.At present,molecular research on A.germari only focuses on gene identification and expression profiling,and there is no report on the functional study of responsing information compounds in A.germari.In this study,we sequenced and compared the antennae,tarsal segment of forefoot,mouthparts(maxillary palps and labial palps),and abdominal end transcriptome of A.germari.We excavated olfactory related genes and analyzed the expression level of these genes.Then we further investigated the binding ability of odorant binding proteins which antennae highly express to pheromones and host volatiles.We used fluorescent competitive binding technique to examine the binding properties of these proteins and confirmed their interactions through molecular docking.(1)After sequencing and analyzing the transcriptome of antenna,tarsal segment of forefoot,mouthparts and the end of abdomen of A.germari,50813 Unigenes were obtained through assembly.Homology and structural domain comparison was conducted on the olfactory genes annotated by NR,and a total of 236 olfactory related genes were identified,including 92 OBPs,64 ORs,34 CSPs,27 GRs,10 IRs,7 SNMPs.(2)We analyzed the phylogenetic tree of olfactory related genes and investigated the differences in expression levels of some olfactory genes in male and female antennae,tarsal segment of forefoot,mouthparts,and abdominal ends of the A.germari.The results of the OBP study in A.germari showed that five AgerOBPs clustered on the PBP branch,with AgerOBP10 highly expressed in the abdominal terminal and AgerOBP27,AgerOBP61,AgerOBP67 and AgerOBP70 specifically expressed in the antennae;the results of the OR study in A.germari showed that AgerOR2,AgerOR3,AgerOR5,AgerOR13,AgerOR33,AgerOR34,AgerOR36,AgerOR40 and AgerOR48 were clustered with sex pheromone receptors and were specifically expressed in male antennae;the results of the study on IR of mulberry aspen showed that AgerIR3 and AgerIR9 were clustered in IR25 a and IR8 a,respectively.The tissue expression profile analysis of CSP showed that most of the AgerCSP genes were widely distributed in the tissues and might be involved in other physiological life activities of the insects besides olfactory recognition.(3)In this study,the prokaryotic expression of highly expressed AgerOBP23,AgerOBP29,and AgerOBP63 in antennae was carried out.First,the binding ability of AgerOBP23,AgerOBP29,and AgerOBP63 to 44 plant volatiles and 14 other sex pheromone of longicorn beetles was studied by fluorescence competitive binding experiment.The results were as follows: AgerOBP29 and AgerOBP63 were narrow,and had no obvious binding ability to the measured odor substances.The binding spectrum of AgerOBP23 was relatively wide,it is associated with the longicorn beetle sex pheromone Fuscuml acetate and eight plant volatiles,Farnesol,cis-3-Hexenal,Nerolidol,Myristol acetate,cis-3-Hexenol benzoate,(-)-α-Cedarene,3-Ethylacetophenone and Decane have strong binding capacity.Homology modeling and molecular docking clarified the key amino acid sites in the binding of AgerOBP23 to different ligands,and to some extent verified the results of fluorescence competitive binding assays.In this study,the olfactory genes were identified by mining transcriptome data from the antennae,tarsal segment of forefoot,mouthparts and abdominal end of A.germar.The relevant olfactory mechanism of A.germar was elucidated by studying the characteristics of OBP in response to specific compounds,providing a reference for the subsequent development of attractants.