The Genetic Mechanism Of Sex Pheromones Variation In Sister Species Helicoverpa Armigera And Helicoverpa Assulta

Reproductive isolation between different species and races of the same species has been a hotspot in the research of species formation.Many moths use some similar chemicals in structure to isolate from each other.Moreover,some sister species may use same chemicals with different ratio to realize reproductive isolation.In recent years,there have been reported on the production and genetics of sex pheromone of moths,which provide further knowledge on the foundation of reproductive isolation.However,there is limited that how sex pheromone production is changed.Helicoverpa armigera and H.assulta are two important agricultural pests of the Lepidoptera Noctuidae family.H.armigera is a highly omnivorous insect,infest nearly 200 species of 20 families of plants.Moreover,H.armigera has strong fecundity and distributes widely,which is hard to control.However,H.assulta is an oligogamous insects,mainly infesting Solanaceae Plants.The larvae and adults of H.armigera and H.assulta,living in the same area,are very similar in morphological structure and are difficult to identify.Theyuse(Z)-11-hexadecenal and(Z)-9-hexadecenal as their sex attractant pheromone components but in opposite ratios.The major sex hormone components of Helicoverpa armigera is cis-11-hexadecenal((Z)-11-hexadecenal,Z11-16:Ald),while the main sex hormone components of H.assulta is cis-9-hexadecenal((Z)-9-hexadecenal,Z9-16:Ald).The ratio of Z11-16:Ald/Z9-16:Ald in Helicoverpa armigera and Helicoverpa assulta reversed as reported.Under natural conditions,the two kinds of sex pheromones are released by different proportion to attract males to mate and realize reproductive isolation.Under laboratory conditions,the two insects can cross and produce fertile progeny.In the laboratory,we are going to let H.assulta cross with H.armigera by single mating,then let hybrids backcross to H.armigera and H.assulta,respectively,by single mating,too.After the phenotype of the ratio of sex pheromone is determined,the individual is used for molecular analysis by AFLP,cloning and sequence,to understand the molecular basis for sex pheromone ratio variation of these two sister species.First,we cut sex blend of H.armigera and H.assulta.The sex pheromone components,Z11-16:Ald and Z9-16:Ald,were measured using GC-MS by ion selection method.The ratio of Z11-16:Ald/Z9-16:Ald was 96:4 for H.armigera female moths and the ratio of Z11-16:Ald/Z9-16:Aid was 4:96 for H.assulta,conformed previous study and provided firmed foundation for the current study.Then,H.armigera and H.assulta were crossed,the phenotype of sex pheromone was determined by GC-MS and individuals belonging to the phenotype of H.armigera and H.assulta,respectively,were counted to determine the genetics of sex pheromone ratio reversed of H.arimigera and H.assulta.When H.assulta(S)female crossed to H.armigera(R)male(SR),it produce fertile both sex offspring.SR female mainly produced Z1-16:Ald,which is like the phenotype of H.armigera.SR female backcrossed to H.armigera(R)male(BC-SRR)that all BC-SRR female mainly produced Z11-16:Ald,being likeH.armigera.However,SR female backcrossed to H.assulta(S)male(BC-SRS,half BC-SRS female mainly produced Z11-16:Ald(being likeH.armigera),and half BC-SRS female mainly produced Z9-16:Ald(being likeH.assulta).Similarly,H.armigera(R)female crossed to H.assulta(S)male(RS)produced fertile male.When RS male backcrossed to H.armigera(R)female(BC-RSR),all BC-RSR females mainly produced Z11-16:Ald,being like the phenotype of H.armigera.When RS male backcrossed to H.assulta(S)female(BC-RSS),half BC-RSS female mainly produced Z11-16:Ald(being like H.armigera),and half BC-RSS female mainly produced Z9-16:Ald(being like H.assulta).It is inferred that the sex pheromone ratio of the two insects is controlled by a single gene.Finally,we used DNA-AFLP to map grandparents,parents,and backcross family females to locate gene corresponding for sex pheromone ratio reversed of H.armigera and H.assulta.We extracted DNA,cut,ligate,amplify,selective amplify,run gels.We obtained 5 markers,combined pheromone phenotype of each moth.Through gel cutting and sequencing,the nucleotide sequence of each marker was obtained,which provide basis for late gene location and identification.In this study,we used single pair mating to get hybrid and backcross family to analyze the proportion of sex pheromone components in parental,hybrid and backcrossed females by GC-MS.The result confirmed the genetics of sex pheromone reversed ratio of the two sister species is controlled by single gene on auto-chromosome,we also obtained 5 molecular markers by DNA-AFLP and explored the genetic mechanism of sex pheromone reversed ratio of H.armigera and H.assulta.The study provides firmly basis for further gene mapping and may show lights on new ideals and new techniques for pest management.