Molecular Characteristics Research Of18S Ribosomal DNA And28S Ribosomal DNA Of Longicorn Beetles(Coleptera: Cerambycidae)

Research on higher taxa of Cerambycidae was traditionally based onmorphology and paleogeography paleontology, however some disputes were stillremained from different views. Nowadays molecular phylogeney study on higher taxaof Cerambycidae remains limitied, therefore we chose18S rDNA and28S rDNA asthe molecular marker to reconstruct the higher taxa of Cerambycidae.Partial sequence of18S rDNA(V4, V7) and28S rDNA (D2, D3) has beensuccessfully amplified which were obtained from63longicorn beetles representing2families and6subfamilies. Molecular data has been used to reconstruct the phylogenyrelationships of Lamiinae, Cerambycinae, Lepturinae, Prioninae, Aseminae, Philinae,Disteniidae and Vesperidae with the morphological and paleogeography paleontologyresults to provide a further elucidating the phylogenic study of Cerambycoidea.In order to explore the evolutionary relationship of higher taxa of Cerambycidae,both18S rDNA(V4, V7) and28S rDNA (D2, D3) has been choosen as the molecularmarker.70species represent6subfamilies and3families refers to the18S rDNAanalysis which include49species were newly sequenced and21species derived fromGenBank database,67species represent6subfamilies and2families refers to the28SrDNA analysis which include62species from this study and5species derived fromGenBank database,63species represent6subfamilies and2families refers to thecombined analysis which were newly sequenced in this study.The main results of sequence analysis by MEGA5.1as follows;(1) In18S rDNA (V4, V7) analysis which include49species were newly sequencedthe Genbank accession number was:18S rDNA V4: KF141946~KF142008and18S rDNA V7: KF142010~KF14207. Base composition of A, T,C, G were21.1%,26.3%,23.6%,28.9%, G+C（52.5%）＞A+T（47.4%）was not obvious.In the18S rDNA variable sites analysis98variable sites and45parsimony informative sites accounting for13.9%and6.4%respectively.(2) In the28S rDNA(D2, D3) analysis which include62species were newlysequenced the Genbank accession number was KF142075~KF142137. Basecomposition of A, T,C, G were18.1%,19.0%,29.6%,33.3%, G+C (62.9%)＞A+T (37.1%) was obvious. In the28S rDNA variable sites analysis331variable sites and279parsimony informative accounting for52.0%and20.0%correspondingly.(3) In the Combined analysis base composition of A,T, C, G were19.7%,23.3%,26.2%,30.8%respectively; G+C (57.0%)＞A+T (43.0%) was obvious; In thevariable sites analysis446variable sites and279parsimony informative sitesaccounting for32.0%and20.0%. Both18S rDNA(V4, V7) and28S rDNA(D2,D3) contains variable regions and conservative regions,28S rDNA(D2, D3) wasmore variable.(4) Base substitution analysis: Transitions (Ts) was the main form of base mutationswhich primarily occurred between T–C, the Transition(Ts) and Transversion(Tv)ratio reached2.76,1.85,1.73in the18S rDNA,28S rDNA and combinedanalysis separately. Nucleotide substitution saturate analysis showed combinedanalysis has the better linear relationship than the single gene analysis whichmeans base mutations was not reached saturate.(5) Uncorrected P-distance analysis: Uncorrected P-distance has been computed byMEGA5.1between subfamilies in18S rDNA(V4, V7),28S rDNA(D2, D3) andcombined analysis separately, similar results has been showed as follows:Philinae and Disteniidae has the maximum distance compared with thesubfamilies of Cerambycidae, Lamiinae has the minimum distance withCerambycinae, Aseminae was close to Lepturinae and Disteniidae,Cerambycinae was close to Prioninae; Some divergence also existed as follows:Vesperidae was close to Philinae in the18S rDNA analysis and Philinae wasclose to Prioninae in the28S rDNA analysis.(6) Phylogeny tree analysis. Phylogeny tree of18S rDNA,28S rDNA and combinedanalysis has been reconstructed with the method of neighbor Jointing (NJ),maximum likelihood(ML) and Bayesian inference(BI) separately. Similartopologies has been showed as follows: Disteniidae and Aseminae divergedearliest in the phylogeny trees, Cerambycinae Prioninae Lepturinae and Philinaein the same branch, Philinae diverged earliest then the divergency order was: Lepturinae→Prioninae→Cerambycinae, Prioninae was close toCerambycinae. Lamiinae was monophyletic group on the top of the phylogenytrees. Vesperidae was close to Philinae than Cerambycidae in the18S rDNAanalysis, Prioninae was close to Cerambycinae, Lamiinae was not the oldestgroup in this study, Philinae should consider as the subfamily level, besides this,the taxonomic status of Lepturinae was not clearly defined in this study, but theresults showed Lepturinae was not the oldest group, it was placed betweenAseminae and Cerambycinae.