Regulate Lifespan Gene Methuselah-like And RNAi In Dastarcus Helophoroides

Dastarcus helophoroides(Fairmaire)(Coleoptera:Bothrideridae),the larvae parasitize various forest trunkborer-insects of larvae and pupae,It is the most effective natural enemy of large-body longhorned beetles,buprestid beetles and snout beetles in biological control,Dastarcus helophoroides adult has relatively longer lifespan.In the study of the Methuselah-like(mth-like/mthl)is connected with lifespan,resistance,and aging mechanism in Dastarcus helophoroides.but it is still unknown the function of mth-like genes receptor proteins in Dastarcus helophoroides,the aim of this study investigate the function of mth-like genes receptor proteins,To explore the function and expression of mth-like genes,establish a RNAi technique using microinjection system in Dastarcus helophoroides.the main research contents and results summarized as follows:1.Three cDNA clones of mth-like genes with full length sequences were obtained from the D.Helophoroides transcriptome database.mth-like11,mth-like2-4,mth-like2-5,and the login Numbers are MH175212,MH175213 and MH175214 respectively.Online prediction and phylogenetic analysis showed that Mth-likes have conserved seven transmembrane domains,Mth-like 11 ? Mth-like2-4 and Mth-like2-5 were more closely evolutionary relationships with similar genes in Tribolium castaneum.2.The mth-like2-2 expression quantity highest in the larval stage,in the pupal stage,the expression of mth-like2-5 quantity highest,mth-like2-5 in the adult just feather express quantity is significantly higher than other mth-like genes,mth-like2-2 was interfered in the larval stage,and mth-like2-5 was interfered in the adult stage.3.Larva injection of 0.3ug/uL,1.2ug/uL and 4.8ug/uL dsRNA could inhibit the expression of mth-like 2-2.The expression of low-dose treatment group was only 41.7% of the control group injected with dsGFP,and the inhibition effect was significant in the medium dose and high-dose treatment group.However,the high dose treatment group had a low rate of cocoon malformation,pupal formation and a high mortality rate.4.Injected 0.3ug/uL,1.2ug/uL and 4.8ug/uL dsRNA could inhibited the expression of mth-like2-5 in Dastarcus helophoroides adult,and the higher the concentration,the lower the expression The high expression of stress was obvious in the low-dose treatment group,the high expression of stress was weak in the middle-dose treatment group,The high-dose treatment group showed little stress hyperexpression.With the increase of dsRNA concentration,the weight of male and female insects decreased and their mortality increased,the spawning time of females in the treatment group of 0.3ug/uL and 1.2ug/uL was delayed,and females in the treatment group of 4.8ug/uL were thrown unspawning until the 90 th day.5.The stress response of mth-like in Dastarcus helophoroides adult: after gene RNAi under hunger,high temperature and oxidation stress,1.2 ug/uL target gene expression of dsRNA treatment group were significantly lower than the negative control group.High temperature and oxidative stress significantly reduced both male and female expression of target genes,the high stress expression of female under starvation stress reaction obviously,the relative expression of male decrease continuously.