Determination And Analysis Of Mitochondrial Genomes Of Four Species Of Insects

Coleoptera also known as beetles,composed of four suborders Archostemata,Myxophaga,Adephaga and Polyphaga.Cucujiformia contains about 230000 kinds,composed of Cleroidea,Lymexyloidea,Cucujoidea,Tenebrionoidea,Chrysomeloidea and Curculionoidea.Many species are seriously harmful to animal husbandry,agroforestry and storage industry,which research more extensive in Coleoptera.The four species of this study are all forest pests,their damage to forest is huge,our country has become one of the countries most suffering from forest pests.With the continuous development of molecular biology,in recent years,mtDNA has been widely used as molecular markers and in the fields of insect phylogeny,genetic variation and structure of populations,identification of related species,etc.In this research,we used High throughput sequencing combined with traditional PCR amplification successfully sequenced mitochondrial genomes of Euplatypus parallelus,Euplatypus solutes,Dendroctonus armandi,Scolytoplatypus Mikado,the new mitochondrial genomes were annotated with reference to the mitochondrial genome informations of other Curculionoidea beetles,Combining with other Cucujiformia mitochondrial genomes in the database,phylogeny of Cucujiformia has been reconstructed.The main conclusions drawn from this study were listed below:1.The obtained sequences length of E.parallelus,E.solutes,D.armandi,S.Mikado,mitochondrial genomes are 16096bp,16117bp,1610 1bp,16438bp respectively.E.parallelus including 22tRNAs,E.solutes including 23 tRNAs,D.armandi and S.Mikado including 21 tRNAs.Four species contain 13 protein coding genes,2 rRNA and a control region.The results are consistent with the mitochondrial structure of the published Cucujiformia.2.The mitochondrial genome of the four study species obvious have an AT skew,whose A+T content can be as high as 75%.All mitochondrial genome have intergenic region and overlapping region,and a conserved motif(ATAA)in overlapping region between the ATP8 and ATP6 genes can be found in all four species.In addition,one conserved motif(TTAT)can be found in the overlapping sequence between ND4 and ND4L genes.3.The functional RNA of the four species included tRNAs and rRNAs.Nearly all of the tRNAs could form the classical clover leaf structure,while tRNASer(AGN)was excluded for its DHU arm was lost.There were many mismatches in tRNAs secondary structure,and in the presence of mismatches,G-U mismatch was the main type.The tRNAIle of the E.parallelus and E.solutes,was located in the control region of the mitochondrial genome.Most of the anticodons of the tRNAs were identical and completely conserved,while the tRNAIle of E.parallelus,E.solutess used the atypical AAU as theiranticodon.not use the normal anticodon GAU.4.All mitochonerial(?)nggenes of 52 kinds of Cucujiformia beetles in this study have an obvlous AT skew,The statistics of start codon use of the mitochondrial protein-coding genes in Cucujifonnia,beetles show that most of the genes used ATG as their start codon,while ATA,ATT and ATC also occupy a certain proportion.No matter Protein-coding genes or single protein-coding genes highest frequency use of Amino acid is the Leu,use frequency is 15.89.5.The phylogeny of Cucujiformia was reconstructed using ML and BI method based on all 13 protein-coding genes.The monophyly of Curculionoidea,Tenebrionoidea.Cleroidea,Chrysomelidae were supported in the analysis.The Euplatypus parallelus and Euplatypus solutes belong to the same genus clustered into one branch then clustered with Scolytoplatypus Mikado,at last clustered with Dendroctonus armandi.So there is a closed relationship between them.Xylotrechus grayii,Monochamus alternates,Obrium sp.Of Cerambycidae,Spiniphilus spinicornis of Vesperidae Separate with Chrysomeloideain in ML and BI.The monophyly of Chrysomeloidea and Cucujoidea were not supported in the analysis.