Genetic similarities and phylogenetic analysis of wild and domesticated species of sheep based on mitochondrial genome

Mitochondrial DNA (mitogenome) is a small and extra-chromosomal DNA, located in the cytoplasm and presents an ideal model to study evolution and genetic similarity. Classical phylogenetics is based on the morphological characteristics of the organisms, while in modern approaches, the phylogenetic distance and its related comparative methods are estimated based on observed genetic diversity in the studied genetic sequences. The aim of the present study was to investigate the divergence and percentage of genetic similarity along with the phylogenetic analysis of the seven main known species of wild and domestic sheep based on the complete sequence of the mitochondrial genome and the separate sequences of 13 protein-coding genes for each genome.
In the present study, complete mitochondrial genome sequences along with separate sequences of 13 protein-coding genes (including NADH ubiquinone oxidoreductase (ND1, ND2, ND3, ND4, ND5, and ND6), cytochrome c oxidase (COX1, COX2, and COX3), ATP synthase (ATP6 and ATP8), NADH dehydrogenase 4 L (ND4L), and cytochrome b (CYTB))  per each genome from six wild species of sheep including Asian Mouflon (Ovis orientalis), Bighorn (Ovis canadensis), Argali (Ovis ammon), Urial (Ovis vignei), Snow (Ovis nivicola), Dall (Ovis dalli), and domesticated species of sheep (Ovis aries) were retrieved from NCBI database and compared to each other. Mitochondrial genomes and genes’ alignment were accomplished by the MegAlign module of DNASTAR software and compared by the Clustal W method. The Sequence Distances sub-section of the MegAlign module of DNASTAR also was used for the analysis of complete genome and gene sequences divergence and similarity percentage. For phylogenetic analysis, complete mitochondrial genomes and protein-coding genes’ sequences were aligned using MEGA7 software. Based on the alignment, phylogenetic tree was constructed using the maximum likelihood method. The percentage of replicate trees of 1000 replicates of bootstrap test was used to represent the evolutionary history for the studied sheep species.
The results obtained from sequence distance analysis showed high genetic similarity (99.8 %) between Ovis orientalis and Ovis aries. Also, the lowest similarity (95 %) was observed between Ovis aries and Ovis dalli. In phylogenetic analysis, two main clusters, each with different sub-clusters were identified. Domesticated species of sheep (Ovis aries) along with Ovis orientalis, Ovis vignei, and Ovis ammon wild species of sheep formed distinct cluster, and Ovis nivicola, Ovis dalli, and Ovis Canadensis fell in a same cluster. In terms of all 13 protein-coding genes (including NADH ubiquinone oxidoreductase (ND1, ND2, ND3, ND4, ND5, and ND6), cytochrome c oxidase (COX1, COX2, and COX3), ATP synthase (ATP6 and ATP8), NADH dehydrogenase 4 L (ND4L) and cytochrome b (CYTB)), the results obtained from sequence distance similarity analysis and phylogenetic trees were similar to the sequences of the complete mitochondrial genomes. More than 99% of the genetic similarity for ND1, ND2, ND5, ND6, COX1, COX2, COX3, and ATP6 genes and 100% of the genetic similarity for ND3, ND4, ND4L, ATP8, and CYTB genes between domestic sheep (Ovis aries) and Mouflon (Ovis orientalis) sheep species were found. Also, similar to the results obtained from the comparison of the complete mitochondrial genome, the domestic sheep species (Ovis aries) showed the least genetic similarity with the Dall (Ovis dalli) and Bighorn (Ovis canadensis) wild sheep species in all 13 protein-coding genes. Similar to the results of phylogenetic analysis of complete mitochondrial genomes, domestic sheep species (Ovis aries) together with Ovies orientalis wild sheep were placed in the same sub-cluster and Ovis vignei and Ovis ammon species were placed in other distinct sub-clusters. In addition, the Ovis nivicola, Ovis dalli, and Ovis canadensis wild species fell in another main cluster, and in this cluster, Ovis dalli and Ovis canadensis were placed in a similar sub-cluster and Ovis nivicola in another distinct sub-cluster.
In previous studies, small parts of the mitochondrial genome (such as a part of the control region or cytochrome b gene) have been considered to show the genetic differences and phylogenetic relationships between different species and breeds of sheep. In the present study, the complete sequences of the mitochondrial genome along with the complete sequences of 13 protein-encoding genes per each genome in seven main species of wild and domestic sheep have been considered for examining genetic similarity and divergence and phylogenetic analysis for the first time. Based on the results obtained from the present study, mitochondrial genome sequences could be used for accurate phylogenetic analysis and clustering of different species of sheep.