Sequencing And Analysis Of The Complete Motochondrial Genome And Segments Of Y Chromosome Of Microtus Fortis

Microtus fortis is classified as Microtus, Arvicolidae, Cricetidae, Rodentia, Mammalia on taxonomy, and mainly distributes among more than 17 provinces or autonomous regions in China and some areas of Russia, North Korea, Mongolia and other places. Since the 1970s, Vole Outbreaks in Dongting Lake has occurred several times, causing huge losses of Agriculture and Forestry, leptospirosis, epidemic hemorrhagic fever and other diseases. Meanwhile, Microtus fortis is also an important experimental animal resources, and has been used to establish many disease models, such as Anti-schistosomiasis, Diabetes and Ovarian cancer. It's obvious that we should study the population genetic structure of Microtus fortis firstly, in order to further research its biological control and use it as experimental resources much better.Mitochondrial DNA and Y chromosome are both used as effective tools to explore the evolutional history from maternal and paternal aspects. Microtus, involving over 60 extant species, is one of the most excellent materials used to do evolutional researches. In recent years, mtDNA has been widely used as a useful marker system in phylogenetic analysis, and there are only two complete mitochondrial DNA genomes, which have been sequenced in Microtus. Y-chromosomal markers have been applied in human population genetic studies, whereas genetic data tracing paternal lineages are still rare in other mammals.The complete mitochondrial genome and some segments of ChrY for Microtus fortis calamorum have been sequenced successfully in this study. They were amplified by methods of the conventional, long distance PCR and walking sequencing of primers. The results follows:1. Mitochondrial genome DNA of M. f. calamorum is a closed-circular molecule, with the length of 16310bp, including 37 genes and control region, the same with other vertebrates.2. Most frequently used initiation codons for 13 protein-coding genes in M. f. calamorum is ATG (COX1, COX2, ATP8, ATP6, COX3, ND4L, ND4, ND6, Cytb), followed by the initiation codon, ATT (ND3 and ND5), GTG(ND1), and ATA (ND2). When it came to stop codons, the coding regions of the gene Cox1, Cox2, ATPase8, ATPase6, ND3, ND4L, ND5, and Cyt b were terminated with TAA, while that of ND2 and ND6 was TAG. An incomplete stop codon T(--) was found in the gene ND1,Cox3 and ND4.3.21 out of 22 tRNA genes could fold into a clover-leaf secondary structure.The tRNAser(AGY) was only 59 bp in length without the dihydrouridine stem loop, which is found in many other rodent animals.4. The internal organization of the CR of M. f.calamorum showed extended termination associated sequences(ETAS-1 and ETAS-2), central domain and conserved sequence blocks 1 (CSB-1). But we could not find CSB-2 or CSB-3 in M. f.calamorum, just as another two species of Microtus reported in other papers.5. The putative origin of replication for the light strand (OL) of M.f.calamorum showed the high conservation in the stem and adjacent sequences. The adjacent sequence 5'-TAAGG-3'of OL stem was found in all three genus Microtus species. However, the adjacent sequence of OL stem is 5'-GCCGG-3'in other mammals.6. In order to investigate the phylogenetic position of M.f. calamorum, the phylogenetic trees (Maximum likelihood and Bayesian methods) were constructed based on 12 protein-coding genes (except for ND6 gene) on H strand from 16 rodent species. M. f. calamorum was classified into genus Microtus. Arvcicolinae. Further phylogenetic analysis results based on the cytochrome b gene ranged from M. f. calamorum to one of the subspecies of Microtus fortis, which formed a sister group of Microtus. middendorfii in the genus Microtus.7. We obtained sequences about 7300bp long on ChrY of M. f. calamorum and M. f. fortis respectively, for the first time, and found 4 SNPs by comparing the sequences of these two subspecies.In this study, we sequenced the complete mitochondrial genome and segments of Y chromosome of M. f. calamorum, which would lay a solid foundation for exploring the maternal and paternal genetic structure of M. f. calamorum. Furthermore, we proposed a new strategy to obtain the sequences of Y chromosome.