| The Hyphantria cunea（Drury）is a quarantine pest of agriculture and forestry in my country.At present,the long-term use of chemical pesticides in China to control the white moth has led to drug resistance and many environmental problems.Therefore,it is very necessary to study the green control methods of H.cunea.Combined with the reported key genes CYP4Gs involved in CHC synthesis in the previous study of this study,the gene sequences of HcCYP4G1,HcCYP4G2 and HcCYP4G3 key genes of epidermal hydrocarbon synthase were obtained by screening from the transcriptome database of H.cunea.In this experiment,the spatiotemporal expression pattern of HcCYP4Gs gene in the larvae of H.cunea was explored,and the role of HcCYP4Gs gene in the drought tolerance physiology of H.cunea was clarified by RNAi technology and drying stress bioassay.To provide exploratory research for pest control methods based on the inhibition of insect CHC synthesis,and to provide theoretical basis for the development of biological preparations of H.cunea.The research conclusions are as follows:1.The results of bioinformatics analysis of the HcCYP4Gs gene of H.cunea showed that the amino acid sequence of HcCYP4Gs had the highest similarity with the A.plantaginis,and the amino acid sequence similarity with the Lepidoptera Spodoptera insects,and Low amino acid sequence similarity with other order insects CYP4G subfamily,and the results were consistent with the morphological taxonomy.The predicted molecular weights of the HcCYP4Gs gene-encoded proteins were 63.03 k Da,64.32 k Da,64.83 k Da,and the p I were8.59,8.06,and 8.31,respectively.The two most abundant amino acids in the polypeptide chain of HcCYP4Gs-encoded proteins were Leu and Lys.All three proteins were unstable and soluble proteins.HcCYP4Gs have no signal peptide,and HcCYP4G1 and HcCYP4G3 have a transmembrane region with 41,57,and 40 potential phosphorylation sites,respectively.2.The study of the spatiotemporal expression of HcCYP4Gs gene in H.cunea showed that there was no significant difference in the expression levels of HcCYP4G1 and HcCYP4G3 at different ages in the 4th instar larvae.The expression of HcCYP4G2 on the 3rd day and 4th day was significantly lower than that on the 1th day and 2nd day.The expression of HcCYP4Gs in 5th instar larvae at different ages was not significantly different.HcCYP4Gs were expressed in all parts and tissues of 5th instar larvae.HcCYP4G1 and HcCYP4G2 were most expressed in the abdomen,and HcCYP4G3 was the highest in the head.The results showed that the expression levels of HcCYP4G1 and HcCYP4G2 in the 4th and 5th instar larvae were much higher than those of HcCYP4G3.3.The results of the RNAi study of the 4th instar of H.cunea showed that after injection of ds HcCYP4G1 in 4th instar larvae,HcCYP4G1 was significantly inhibited on the 1st,2nd and 3rd days.The silencing efficiencies were 69.30%,65.38%and 78.03%on days 1,2 and 3after injection,respectively.And it has significantly reduced the expression level of HcCYP4G1.After injection of ds HcCYP4G2,it has no inhibitory effect on the HcCYP4G2gene after testing,and the specific reasons need to be further studied.The results showed that the 4th instar larvae of H.cunea had a good response to ds HcCYP4G1,and HcCYP4G1 could be a new target for genetic control of H.cunea.4.The results of the study on the drought tolerance of the 4th instar larvae after RNAi showed that the mortality of larvae injected with ds HcCYP4G1 at RH=25%was significantly higher than that of larvae injected with ds GFP and dd H2O,and the mortality of larvae injected with ds HcCYP4G2 was not different from that of larvae injected with ds GFP and dd H2O.The larvae that died after being treated in a dry environment suffered from severe water loss,the body length was significantly smaller than that of the normally grown larvae,and the larvae were obviously shriveled and blackened.The results combined with RNAi also indicated that HcCYP4G1 could serve as a good target for the genetic control of H.cunea. |
