Identication And Functional Research On Odorant Binding Proteins From Liriomyza Trifolii (Burgess)

The leafminer Liriomyza trifolii is a is a worldwide pest of polyphagia,it can harm a variety of vegetables and flowers and result in a decline in yield and quality.Odorant binding protein(OBP)is a kind of acid protein with small molecular weight,it is mainly in the lymphatic fluid of the insect olfactory sensillum and could transfer the chemical signals to the receptor,then causes the olfactory nervous reaction.This study identify and clone the OBPs sequence according to the transcriptome of L.trifolii and the spatiotemporal expression were analyzed and then the recombinant protein was expressed and purified.The characteristics of odor binding and immune localization also were studied.It provides a theoretical basis for the study of the odor binding protein and the olfactory mechanism and provides new ideas and methods for the green control of L.trifolii.We screen and verify the results of the transcriptome,then nine odorant binding proteins of L.trifolii were found and named as LtriOBP1-9,the LtriOBP1-9 were divided into three sub-families: 7 Classic OBPs,1 Minus-C OBP,1 Dimer OBP.Rootless evolutionary trees show low homology among LtriOBP1-9.But LtriOBP1-9 have high homology with Liriomyza sativae OBPs.qPCR was used to analyze the spatiotemporal expression of these genes.In different development stages,the results showed LtriOBP5 highly expressed in the larva stage,LtriOBP4 highly expressed in the pupa stage,the other 7 LtriOBPs highly expressed in the adult stage and the exression level in male adult were higer than female adult.The expression of males increased or decreased within the 0-5 days.In different tissues,the results showed in addition to the specific expression in the antennae,some LtriOBPs also expressed in the head and leg.The differences of expression reflected the LtriOBPs not only have the function to recognite odors in olfactory response,but also take part in the development and gustatuin.In order to study the protein function of LtriOBPs,7 LtriOBPs were prokaryotic expressed and purified.We obtained the LtriOBPs with high purity.1-NPN was used as fluorescent probe to study the binding properties of LtriOBP1-7 with twenty-five different odor ligands.LtriOBP1 could bind to 13 odor compents and has good combining capacity with ?-Ionone,Linoleic Acid.LtriOBP2 could bind to 21 odor compents and has good combining capacity with?-Ionone,1-Octen-3-ol,Benzothiazole.LtriOBP3 could bind to 9 odor compents and has good combining capacity with ?-Ionone.LtriOBP4 could bind to 3 odor compents and has good combining capacity with Benzothiazole,Linalool,4-Ethylacetophenone.LtriOBP5 could bind to 3 odor compents and has good combining capacity with?-Ionone,Linoleic Acid,Linolenic Acid.LtriOBP6 could bind to 8 odor compents and has good combining capacity with Linoleic Acid,Linalool.LtriOBP7 could bind to 21 odor compents and has good combining capacity with ?-Ionone,Linoleic Acid,Linolenic Acid.The binding properties of LtriOBP1-7 provide the reference to exploit the repellent.The frozen section and immunofluorescence staining technique to study the distribution of the LsatOBP1-7 in the antennae,head,mouthpiece,protarsus,larva and pupa of L.trifolii.The distribution of antennae flagellum showed there were some LtriOBPs which could strongly bind many odors in the trichoid sensilla and coeloconica sensilla.And that indicated these two kinds of sensilla could take part in the reaction about recognition of host plants.The LtriOBPs which were located in the basiconic sensilla have high selectivity to odors,that indicated the basiconic sensilla could take part in some specific olfactory.What is more,LtriOBP2,5 were found in the antennae pedicel which explained pedicel also could identify some odors and participate in olfactory reactions.LtriOBP2,5,6 were located in the mouthpiece.The result confirmed its high expression in the head and indicated they participate in gestation.LtriOBP2 also was located in the eyes of L.trifolii and they could take part in the visual pigmentation.LtriOBP2,6,7 also were located in the leg and they could take part in the gestation of recogniting non-volatile substances.LtriOBPs were widely distributed in many different tissues and could have multiple physiological function...