The Complete Mitochondrial Genomes Of The Cistoclemmys Flavomarginata And Cistoclemmys Galbinifrons With Inolecular Phylogenetic Analysis Of The Higned Turtles

As one of Earth's most instantly recognizable life forms, turtles have highly specialized morphological characteristics, with approximately 450 nominate species in the world. Over the past two decades, due to anthropogenic killing and pet trading, the majority populations of the wild hinged turtles have been very rare, and they were classified as extremely endangered turtles by IUCN2006 (except Cuora amboinensis and Cistoclemmys flavomarginata), it is urgent to study its conservation biology and phylogenetic evolution. As far as May of 2010, the complete mt genomes of turtles have been released from GenBank for only 34 species, and there are only 5 hinged turtles. To explore the relationships among genus of the hinged turtles and provide a theoretical basis for the protection of these animals, we determined the complete mitochondrial genome (mtDNA) of Cistoclemmys flavomarginata and Cistoclemmys galbinifrons.The complete nucleotide sequences of the mitochondrial genome (mtDNA) of the two turtles were determined using PCR,LA-PCR and sequencing technology. The size of molecules was 16 721 bp, and 17 244 bp, respectively. The genome organization, gene order, and base composition conformed to that of the typical vertebrate, including 22 tRNAs, 2 rRNAs, 13 protein-coding genes, as well as a control region, revealing the high conservation in turle mitochondrial genome evolution. As most turtle's mtDNA reported an extra A nucleotide was discovered in ND3 gene of the two turtles. We also analysed the control region of five hinged turtles, and the length of CR were from 980 bp to 1 722 bp respectively, all containing the termination associated sequences (TAS) and several conserved sequence blocks (CSB-F, and CSB1-3). A highly variable region at 3'end of CR was identified by comparison with other turtles, containing variable number tandem repeats (VNTRs) or called microsatellites. The motifs of tandem repeats (7 types) are made up of 2-8 nucleotides, and the copy numbers are from 4 to 48 containing A,T and C. Furthermore, the differences of the motifs and the repeat times induced the discrepancies of CR length among different species and among these microsatellites. Compared with the microsatellites motifs of the inter-species, the result may provide important reference for the turtle's systematics research, relationship analysis, and protection of species multiplicity and so on.To review phylogenetic relationships among major turtle lineages and the genus of the hinged turtles, maximum parsimony (MP), maximum–likelihood (ML) and Bayesian (BI) analyses were conducted based on concatenated nucleotide sequences of 12 protein- coding genes of mtDNA from 22 turtle taxa and Caiman crocodiles and Alligator mississippiensis as outgroups. The three resultant trees showed homological topologies, and the results strongly supported that Cuora, Cistoclemmys and the monotypic Pyxidea formed a monophyletic Cuora group, so we suggest these three genus should be incorporated into Cuora, whereas Cyclemys was not closer to this clade than to Sacalia, suggesting that Cyclemys and the Cuora group (containing Pyxidea) may have originated from different ancestors; Mauremys appeared to be paraphyletic with respect to Chinemys; Platysternidae is actually the sister to the Emydidae; the Geoemydidae with Testudinidae was a sister group rather than with the Emydidae.