Complete Mitochondrial DNA Sequence Variation Analysis Of Yak And Cattle-Yak Hybrids

China is the largest country in the world with a great number of yaks, and yak resources is very rich. There are five yak breeds included in the Bovine Breeds in China, as much as representative groups of11Chinese yak populations, the genetic diversity has been focused for researchers so far. Blood protein polymorphic, microsatellite markers, the sequences of mitochondrial D-loop region, the cytochrome gene sequences and nuclear gene sequences were analyzed. Main yak genetic diversity in China based on mitochondrial whole genome sequences has not been reported. Yak is an integral part of livestock by the livestock economic development of Qinghai-Tibet Plateau and its surrounding areas, providing meat, milk, wool, draft power, fuels, and other production necessities of life for local herders, but with lower production performance. It can greatly improve the production performance of its milk, meat by cattle-yak hybrid. However, Fl generation male of cattle-yak is sterility. Therefore, to clarify the mechanism of cattle-yak male sterility is significant for yak crossbreeding and heterosis utilization, and improving herdsmen’s living level. Researching on male sterility mechanism in cattle-yak was focused on spermatogenesis meiotic nuclear genes, the research on cattle-yak male sterility by mitochondrion has not been reported.To further study the yak genetic diversity, and the relationship of mitochondrial genome and cattle-yak male sterility, our studying will adopt to clone and sequence and documentation tracking five yak breeds (Tibetan alpine yak, Jiulong yak, Maiwa yak, Qinghai Plateau yak and Tianzhu White yak) in China with36yak individuals and four mitochondrial genomes of cattle-yak. Then we analyzed the genetic composition, sequence features of the Chinese yak mitochondrial genome, the Chinese yak genetic diversity and genetic differentiation, compared the difference between yak and cattle-yak mitochondrial genome. Providing the basis for the yak germplasm characteristics and male sterility molecular mechanisms, we got some main results which was followed:1. Yak mitochondrial genome sequence analysisResearch by cloning and sequencing Tibet Mountain yak in mitochondrial genome sequence, we found that the full-length of yak mitochondrial genome was16324bp, including13protein-coding genes,22tRNA genes,2rRNA genes and an non-coding region. And the gene composition, arrangement and location were basically the same with other species of bovine subfamily. The total length of13yak mitochondrial protein-coding gene sequence was11400bp, there were four kinds of initiation codon. Most protein-coding genes (9) had the ATG start codon, the ND2, ND3and ND5genes had ATA start codon, the ND6gene had the AAT start codon. Yak mitochondrial non-coding region sequence was893bp, located between the tRNA-Pro and tRNA-Phe, containing a termination sequence (ETAS), central conserved region (CD), and conservative sequence District (CSB) compared with other species of bovine subfamily. However, the sequence length of non-coding region among different species of bovine subfamily was species-specific. The total sequences length number of22yak mitochondrial tRNA gene was through66bp to75bp, and the total length was1511bp. Yak mitochondrial genome consists two rRNA genes,12s rRNA and16s rRNA gene, with the957bp and1571bp length respectively.2.Yak mitochondrial genome genetic diversity and genetic differentiation222variable sites were found in36Chinese yak mitochondrial genome sequence, the percentage of polymorphic loci was1.36%. Conversion occurs in the Chinese yak mitochondrial genome sequence as44times, transversions as2times, transition/transversion (Ts/Tv) ratio was18.11, greater than the critical value (2.0) of the transition/transversion, indicating that the Chinese yak mitochondrial genome mutations has not reached saturation.36yak mitochondrial genome sequences can be defined as the30haplotypes, haplotype diversity (Hd) was0.984, nucleotide diversity (Pi) was0.284%, the average number of nucleotide differences (k) was46.3, it showed that the Chinese yak genetic diversity was rich. In five yak breeds, polymorphic loci was through4to170, haplotype diversity (Hd) was through0.833to1.000, nucleotide diversity (Pi) was through0.025to0.486. The average number of nucleotide differences (k) was through4to76.4. Maiwa yak had the highest one in polymorphic loci, haplotype diversity, nucleotide diversity, and the average number of nucleotide differences, indicating that genetic diversity of the Maiwa yak was the richest of five yak breeds in China. Neutrality tests founded that Chinese yak populations and five yak mitochondrial genome sequences correspond to the neutral mutation and population size remained stable, there had no population expansion and sustained growth mode. The genetic distance between Maiwa yak and the other four varieties was the largest. The genetic distance among Chinese five yak groups was about0.002-0.088, with average0.031. Some differentiation was found the varieties of Chinese yak, and the Maiwa yak had the most obvious differentiation between the other yak breeds in China. Chinese yak population genetic different coefficient was0.0242, gene flow was10.08, indicating that the genetic differentiation between Chinese yak breeds obvious, but the most differentiation between Maiwa yak and the other yak breed.3. A comparative analysis of the mitochondrial genome between yak and cat-yak hybridFour cat-yak mitochondrial genome sequences were obtained by cloning and sequencing technology with the length of16339bp-16443bp. And the significant difference was existed between the cat-yak and the yak.784variable sites were founded in the hole mitochondrial genome sequences between the yak and cat-yak. It contained47in non-coding region,619in protein-coding genes,39in tRNA genes,79in rRNA genes. In the non-coding regions, ETAS2was the biggest in the differences, while CSB2, OH, the LSP and HSP were highly conserved in the yak and cat-yak. Compared with the yak mitochondrial non-coding region, cat-yak had the two longer base insertion, which were located in the former ETAS1and between ETAS2adn CD. There were92non-synonymous mutation sites which caused81amino acid residues changed; the type of amino acid residues between the yak and cat-yak were changed of35, which were in92non-synonymous mutation sites, and ATP6protein had the biggest ratio that amino acid residues change as1.76%, but the COI, ND3and ND6protein did not occur to change the type of amino acid residues. In the tRNA genes, differences in the tRNA-Asn gene was the most percent, while the tRNA-Val and other6gene were exactly the same between yak and cat-yak. Secondary structure analysis of variable sites in the D ring, PΨC central, Central loop, receptor arm and anticodon arm, and so on, were analyzed. The results showed that there was1bp insertion or deletion at nt60of tRNA-Asn gene and at nt21of tRNA-Leu (CUN) gene. In the12S of rRNA gene, there were19variable sites in the secondary structure of the loop region and8in the stem region; there were38variable sites in the loop region and14in the stem region in16S rRNA.