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Over the past few years, there has been a significant progress in genetic testing in dogs through the development of genomic maps, which make it possible to determine the background of polygenic genetic diseases. The information obtained by mapping the canine genome, a detailed marker map, and the understanding of genome architecture have changed the possibilities and direction of canine genetic research. It is now possible to understand the genomics of the qualitative and quantitative traits associated with the phenotype, genetic predisposition, and genetic background of canine defects and diseases. Investigations of canine genetic diseases are particularly valuable because their results can be used in human medicine. From the medical point of view, genetic diseases in both humans and dogs are similar, which makes it possible to test new therapeutic approaches in the case of orthologous genes containing mutations responsible for genetic disorders.
The aim of the study was to identify polymorphisms and mutations in the mitochondrial ND4 gene and to analyse the associations between the occurrence of molecular changes in mtDNA and phenotypic traits in tumours in German Shepherd dogs. Fifty samples obtained from blood and tumour tissues of German Shepherd dogs with diagnosed tumours were analysed. DNA extraction, amplification, and sequencing of the mtDNA ND4 gene, and bioinformatics, statistical, and in silico protein coding SNP analyses were performed. ND4 mutations and/or polymorphisms were noted in eleven nucleotide positions in nearly half of the examined dogs. All the changes were substitution mutations. A majority of the changes identified were homoplasmic. In one dog with osteosarcoma, blood heteroplasmy was detected. In two positions of the ND4 gene, presence of non-synonymous mutations leading to amino acid changes in the ND4 protein was reported. Analyses carried out to determine the deleterious effect of mutations indicated an almost 97 and 62% probability that a single amino acid substitution (p.G239V and p.I401T, respectively) in the protein has a negative impact on its function. The results of statistical analyses indicate a significant association between the occurrence of mutations in three loci of the ND4 gene and the location of tumours. The mutations identified may be a result of cell adaptation to the changes in the environment occurring during carcinogenesis. The high frequency of mutations in the tumours may indicate genetic instability of mtDNA, which may also play a role in carcinogenesis.
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