Studies On Cuticular Wax Synthesis Pathway And Related Genes In Cotton Mealybug,Phenacoccus Solenopsis Tinsley

The cotton mealybug,Phenacoccus solenopsis Tinsley(Hemiptera: Pseudococcidae),is an important invasive pest around the world,which infests diverse crops and horticultural plants.In China,it was first found in Guangzhou in 2008 and so far has spread to 124 counties(districts)of 13 provinces posing great threat to agriculture.The body of P.solenopsis female is covered by wax,from which this mealybug benifits for water proofing,preventing desiccation and avoiding predators.However,little is known about the synthesis and related genes of cuticular wax of P.solenopsis.Here,I got some results:(1)I sequenced transcriptome of the cuticle and other tissues of P.solenopsis adult female and got 38506 transcripts.KEGG annotation showed there were fatty acid biosynthesis pathway,fatty acid elongation pathway,terpenoid backone biosynthesis pathway and fatty-acyl reductase(FAR)genes.Blastx was used to search P.solenopsis transcriptome for insect CYP4 G proteins,which related to decarbonylation reaction in insect cuticlular hydrocarbon synthesis,and I found 214 transcripts,which indicating that CYP4 G genes were present in P.solenopsis.These pathways and genes might be part of cuticular wax synthesis pathway of P.solenopsis.(2)Differential expression analysis was performed to find differences of transcriptomes between cuticle and other tissues.I found 342 cuticle up-regulated genes,and related 1172 transcripts from the 367 differential expressed genes and 1272 transcripts.The 1172 cuticle up-regulated transcripts were then annotated by KEGG and GO,and I found 3 fatty acid synthase(FAS)genes,3 elongation of very long-chain fatty acid(ELOVL)genes and 1 FAR gene.These genes were supposed to be highly related to cuticular wax synthesis of P.solenopsis.(3)I sequenced proteome of the cuticle of P.solenopsis adult females and got 5812 proteins.KEGG annotation also showed there were fatty acid biosynthesis pathway,fatty acid elongation pathway,terpenoid backone biosynthesis pathway and FAR genes in proteome.Blastp was used to search P.solenopsis proteome for insect CYP4 G proteins and I found 45 proteins.This suggests the presence of CYP4 G enzymes in P.solenopsis.All these indicate that cuticle of P.solenopsis has the potential ability of synthesizing wax.(4)Blastx was used to search transcripts of three FAS genes,three ELOVL genes and one FAR gene found from differential expression analysis for P.solenopsis protein sequences.All of the 27 transcripts of these genes were aligned to proteins,suggesting that these genes are translated to proteins and they play biological function in the cuticle.(5)I observed wax synthesis of P.solenopsis female 5,10 and 15 days after eclosion.Wax was found to occur faster at 5 and 10 days after eclosion than 15 days.Moreover,the cuticular wax could recover in 3 days after being removed.(6)RNAi was performed for FAS,ELOVL and FAR genes found from differential expression analysis and these genes of high expression in cuticle.During each experiment,I mixed the dsRNA of 2-3 genes of them.Seven of 22 genes were significantly knocked down.Wax synthesis of P.solenopsis was observed for 3 days to find the consequence of dsRNA injection,however,no difference were observed compared to the individuals injected with dsGFP(control).