Functional Research Of Fruit Fly Or82a And Locust LmigOR3

One of Drosophila rmelanogaster OR and one of Locusta migratoria OR are selected to preliminarily explore their characteristics and biological functions in this thesis.?.The function of D.melanogaster Or82aThe results of single sensillum recordings(SSR)showed that Or82a could be strongly activated by geranyl acetone,one of fly body volatiles,even at the low concentration of 0.1%.This response completely lost when Or82a was knocked out and this loss was recovered when Or82a-was rescued.When Or8 2a was knocked out,courtship index for mutant male paired with wild-type female significantly increased compared to the index for wild-type male paired with wild-type female.When Or82a-was rescued,courtship index decreased to normal level of wild type.There was no significant difference between courtship index for mutant male paired with wild-type female in the presence of geranyl acetone of 1:5,000 dilutions and the index for wild-type male paired with wild-type female without the presence of geranyl acetone.Based on the above results,we can draw the conclusions that Or82a can detect geranyl acetone and that geranyl acetone at low concentration can inhibit male courtship index.?.Functional research of LmigOR3 based on Drosophila transgenic technologyFour candidate OR genes were identified from normal cDNA library of fourth instar locust larvae antennae,named LmigOR3,LmigOR4,LmigOR5 and LmigOR6 respectively.LmigOR3 and LmigOR4 were complete genes and the sequences were 1,242 bp and 1,212 bp respectively,which encoded 413 and 403 amino acids respectively.Transmembrane region prediction showed LmigOR3 and LmigOR4 both had seven transmembrane regions,so they were typical insect ORs.LmigOR5 and LmigOR6 were partial genes,in which the upstream sequences were not complete.5' RACE method was used to get the lost sequences.The results showed that the complete sequences of LmigOR5 and LmigOR6 were 1,227 bp and 1,236 bp respectively,which encoded 408 and 411 amino acids respectively.LmigOR5 and LmigOR6 also had seven transmembrane regions,so they were also typical insect ORs.RT-PCR results showed LmigOR3,LmigOR4,LmigOR5 and LmigOR6 were all specifically expressed in antennae,not in tarsi,wings,mouthparts and mid-guts.RNA in situ hybridization showed that LmigOR3 and LmigOR4 were expressed in the neurons housed in trichoid sensilla and LmigOR5 and LmigOR6 in the neurons housed in basiconic sensilla.UAS-LmigOR3 was constructed by inserting LmigOR3 into pUAST.It was injected into the egg of Drosophila mutant Or67dGAL4 and got transgenic fly UAS-LmigOR3 by picking out red-eyed offspring.LmigOR3 was specifically expressed in the neuron housed in T1 sensillum in the transgenic fly.Single sensillum recordings were conducted to record the coding of LmigOR3 for 185 odorants.The results showed that 55 odorants could elicit responses and among these odorants 18 odorants could elicit strong responses(Aspikes/s ? 20)for LmigOR3 at the concentration of 10%(vol/vol or wt/vol),which mainly belonged to ketones,esters and heterocyclics.3-heptanone was the strongest ligand for LmigOR3.Based on the above results,we can draw the conclusion LmigOR3 is a broadly tuned odorant receptor expressed in the neuron housed in Locusta migratoria trichoid sensillum.It is different from the results from flies,moths and mosquitoes,which indicates the locust has own specific pattern of odor coding.