High - Throughput Sequencing Of The All - Round Mitochondrial Genome Of Dragonfly And Analysis Of The Development

The sequence and structure of mitochondrial DNA is a hotspot of animal genome research. Research content include:determination of DNA sequences, analysis of base composition, codon usage, RNA secondary structure, gene organization, and so on. These informations can be used for the research of animal phylogenetic and evolution. Although lots of the whole mitochondrial genome sequences have been reported, but there is only 11 species of the odonata whole mitochondrial sequences have been reported, none in Platycnemididae, and only one species had complete mitochondrial genome sequencing in Calopterygidae, with few relative studies.This study employed the method of High throughput sequencing to determine the whole mitochondrial sequences of Platycnemis foliacea (Platycnemididae) and Agrion atratum (Calopterygidae), the splicing, annotating and analyzing the whole mitochondrial genome were also performed. Then combined the three data sets of PCGs, rRNAs, and rRNA & PCG of the mitochondrial genomes of 11 odonata insects which have been included in the NCBI, with Ephemera orientalis (Ephemeroptera) as outgroup, using Maximum parsimony, Maximum likelihood and Bayesian inference method, the phylogenetic tree were constructed, and the phylogenetic relationship of odonata species were analyzed. Main conclusions are observed and described as followed:1. The determined mitochondrial genome sequeces of Platycnemis foliacea and Agrion atratum are 15418 bp and 15506 bp respectively, including 13 protein genes,2 rRNA genes, 22 tRNA genes and an A+T-rich region. Among these genes,4 protein genes,8 tRNA and 2 rRNA genes encoded by L chain, the rest of the other genes are encoded by H chain. Platycnemis foliacea has 14 mtDNA genetic overlap regions with the total up length of 44 bp; 8 Gene spacers with the total up length of 72 bp. Agrion atratum has 12 mtDNA genetic overlap regions (40 bp in size),10 genetic intervals (77 bp in size). The whole mtDNA sequence, and the base compositions of each part in Platycnemis foliacea and Agrion atratum showed AT bias, A+T content is greater than the G+C content. When the AT and GC skew value are positive, suggesting that the bases of A content is greater than the content of T, G content is greater than the content of C, on the contrary, the bases of A content is smaller than the content of T, G content is smaller than the content of C.2. Among the 13 protein-coding genes in Platycnemis foliacea and Agrion atratum, six protein-coding genes (ND2, C0Ⅱ, ATP8, C0Ⅲ, ND6 and Cytb) and 8 protein-coding genes (ND2, C0Ⅱ ATP8, C0Ⅲ, ND6, ND3, Cytb and ND1) have the initiation codes ATN, respectively.3 protein-coding genes (ND5, ND4 and ND4L) and 4 protein-coding genes (CO I, ND5, ND4 and ND4L) have the initiation codes AAT, respectively. The rest of other protein-coding genes are initiated by TTN and TTG, respectively. All the 13 protein-coding genes of mitochondrial genome in Platycnemis foliacea and Agrion atratum have two kinds of termination codons:complete termination codon and incomplete termination codon. Both Platycnemis foliacea and Agrion atratum mtDNA encode 22 tRNAs, distributed between rRNAs and PCGs. In the tRNAs secondary structure, there are 37,32 gene base mismatches, respectively and the G-U base mismatch are the most. Platycnemis foliacea and Agrion atratum rRNAs gene (s-rRNA and 1-rRNA) are located between tRNALeu(L2) genes and A+T rich region. The control areas of two Damselflies are located between s-rRNA and tRNAIle, A+T% content were 81.3% and 87.1% respectively, with A+T bias.3. In the component of nucleotides A, T, G, C of the 13 odonata insects whole mitochondrial sequences, the base content A is the highest, while the base content of G is the lowest. Among the use frequency of nucleotides in the protein-coding genes (PCGs), H chain and L chain of PCGs, the base content of T is the highest, by contrast the base content of G and C witnessed the lowest. Both the whole mtDNA sequences and the base composition of each part in 13 species of odonata insects have AT bias. The content of A+T is greater than the content of G+C. On the case of codon usage, PCGs, PCGs-H and PCGs-L in 13 species of odonata insects have specific preferences. RSCU analysis suggests that the occurrences of synonymous codons that ended with base A or T is much higher than the rest of the synonymous codon occurrences.4. Using 13 species of odonata insects’ mitochondrial genome PCGs and rRNAs&PCGs data sets to construct the MP, ML and BI phylogenetic trees respectively. In all three trees, Odonata are divided into two branches, and the branches are in a similar situations. In one main branch, Platycnemis foliacea (Platycnemididae) were collected together with Ischnura pumilio (Coenagrionidae), they formed a sister group relationship, Euphaeidae and Calopterygidae and Pseudolestidae were collected together. In another branch, Libellulidae and Corduliidae were collected together. In rRNAs data set, using three methods to construct phylogenetic tree show some differences, when comparing to that of PCGs and rRNAs & PCGs. Phylogenetic analysis results suggested that it is uncertain on the phylogenetic position of Epiophlebiidae. Besides, the relationship between Pseudolestes mirabilis (Pseudolestidae) and Euphaea formosa (Euphaeidae) and the relationship between the three kinds of genus in Libellulidae also remained unclear, further studies are needed.