| Peach aphid(Myzus persicae)is an important prickly pest in fruit tree and vegetable production because of its harmful characteristics of overeating and the spread of plant virus.The aphid propagated rapidly in the growing season by parthenogenesis,and the sexual reproduction was adopted to resist the low temperature environment during overwintering.Therefore,it is very important for peach aphids to complete their life history by identifying sex pheromones to find mate mating.At the same time,peach aphid,as a widely existing phytophagous insect with high density of population density,needs to rely on the perception of alarm pheromone when it is attacked by natural enemies and the population density is too high.Therefore,smell is very important for the survival of aphids.The study of the molecular mechanism of the recognition of pheromone odorant by peach aphid has become a hot field of attention.small molecule odorant transporter odorant binding proteins(odorant binding proteins,OBPs)and chemosensory proteins(chemosensory proteins,CSPs)as well as odorant receptors(odorant receptors,ORs)that recognize odorant substances play an important role in research in this field.In order to further study of peach aphid sex pheromone and molecular and cellular mechanism of alarm pheromones,by using the reverse chemical ecology through regulating the aphids behavior for the purpose of effective prevention and control,this paper first clear the peach aphid tentacles on the smell sensor type,and then through the transcriptome sequencing dug peach aphid smell related genes,and then combining in vitro expression,fluorescence,double electrode voltage clamp and RNAi experiment,the preliminary system has been clear about the OBPs,CSPs and ORs in the recognition of sex pheromone and alarm pheromone function,in the process of the main results were as follows:1.Through scanning and transmission electron microscopy(TEM)observation of the antennae and head of the apterus persicae,it was identified that there were four kinds of olfactive organs on the antennae of the apterus persicae,namely,Primary rhinaria(PR),Trichoid sensilla(TS),coeloconic pegs(CP)and Campaniform sensillum(CS).There are two kinds of trichoid receptors on the head.The results of transmission electron microscopy showed that the neurons in the antennae gradually decreased with the extension to the top,most of which were distributed on the primary sensory circle,and some of which were extended to the trichoid sensilla at the top.These results laid a foundation for the systematic study of the olfactory function of different receptors and the expression and localization of olfactory genes in the receptors.2.The OBPs,CSPs and ORs of peach aphid were systematically identified and analyzed at the genomic and transcriptome levels.The results showed that the number of OBPs/CSPs/ORs in peach aphid was 9,9 and 33,respectively.Sequence ratio and phylogenetic tree analysis were used to determine its nomenclature and evolutionary relationship.3.The function of obp3/7/9 recognition alarm pheromone of peach aphid was systematically studied.In addition to OBP3 3/7,OBP9 is also an important binding protein for the recognition of alarm pheromone(E)-β-farnesene(EβF),and through in vitro expression and binding experiments,the strong binding ability of these three proteins to EβF was confirmed,and the binding constants Kd were OBP3=2.5±0.4μM,OBP7=1.1±0.6μM,OBP9=3.9±0.1μM,respectively.In addition,there is an interaction between these three proteins in the recognition of EβF.In vitro RNAi function determination confirmed that knocking out any one of OBP3 3/7/9 alone showed no significant change in the repellent effect of EβF on peach aphid.Only by simultaneously inhibiting the expression of OBP33/7/9 could the response of peach aphid to EβF be changed at the electrophysiological and behavioral levels.In addition,OBP3 3/7/9 can bind effectively with two pheromone components(4As,7S,7AR)-Schonitolactone and(1R,4As,7S,7AR)-Schonitol.4.The function of detecting alarm pheromone and sex pheromone of peach aphid CSP2/4/5/9 was studied.Through in vitro expression and fluorescence binding experiments,it was confirmed that CSPs could bind sex pheromones and alarm pheromones.The binding constant Kd of CSP4 is 4.21±0.11μm.The binding capacity of CSP5 pairs(4As,7S,7Ar)-Schizonolactone and(1R,4As,7S,7Ar)-Schizonitol was moderate,and the binding constants were 7.42±1.33μm and 7.21±0.75μm.It is suggested that CSP4 is involved in EβF and Csp5 is involved in sex pheromone recognition.5.The odor receptor genes in the peach aphid genome and transcriptome were identified,the gene structure was identified and the function of the gene in vitro was determined.The results showed that a total of 33 ORs were identified,including one atypical olfactory receptor ORco and 32 common ORs.Genetic structure analysis showed that OR20/21/22 was tandem.The OR17/ORco complex was expressed in vitro from Xenopus laevis oocytes,and the recognition of 62 different odors was recorded by the double-electrode voltage clamp technique.The specific binding pheromone components(4as,7S,7a R)-Schizonolactone and(1R,4as,7S,7a R)-Schizonitol were found.It was suggested that OR17 and peach aphid were involved in the identification of sex pheromone components.In this study,by studying the olfactory recognition related genes of peach aphid,the molecular mechanism of its recognition of various pheromones was improved,and the target genes were provided for green prevention and control of peach aphid. |
