Search

Genetic resistance to sperm cryopreservation is an important factor in the sires selection. In our study, we studied the cryoresistance of the sperm in Gallus gallus, Bos taurus, Equus ferus caballus. Sperm quality was assessed using computer-assisted semen analysis (CASA). Genetic studies were performed using Affymetrix Axiom®600k Array chips for cocks, Affymetrix Axiom®600k Array for stallions and BovineSNP50 for bulls. For the analysis of genotypes, the GWAS (Genome-Wide Association Studies) was used. The calculations were carried out using the PLINK and EMMAX programs. The candidate genes found as a result of our work were compared with the functions of orthologous genes, according to the publications of other researchers. As a result of comparing different animal species, a common candidate gene was found, localized on chromosome 3 (Gallus gallus) and on BTA9 (Bos taurus) BCKDHB gene encodes the beta subunit E1 of branched chain keto acid dehydrogenase, which is a multienzyme complex associated with the inner membrane of mitochondria. In roosters, it was associated with the preservation of sperm motility after cryopreservation, and it was found in the analysis of the integrity of the sperm head in bulls. The functioning of genes associated with meiosis and sperm maturation is an important condition for maintaining its quality after freezing. The work of genes responsible for the formation of membranes and various cellular structures associated with the functions of spermatozoa is also an important factor in cryoresistance. Another important condition for the functioning of the sperm is the expression of genes that prepare the sperm for fertilization. These are calcium metabolism genes involved in preparation for capacitation. Changes in the structure of proteins can lead to premature capacitation and disruption of sperm integrity. The found molecular genetic markers will be used to select sires whose sperm are suitable for freezing. Project of RSF № 18-16-00071.

The fertilizing ability of stallion sperm after freezing is lower than in other species. The search for the optimal extender, combination of extenders, and the freezing protocol is relevant. The aim of this study was to compare lactose-chelate-citrate-yolk (LCCY) extender, usually used in Russia, and Steridyl&reg, (Minitube) for freezing sperm of stallions. Steridyl is a concentrated extender medium for freezing ruminant semen. It already contains sterilized egg yolk. Semen was collected from nine stallions, aged from 7 to 12 years old. The total and progressive motility of sperm frozen in Steridyl was significantly higher than in semen frozen in LCCY. The number of spermatozoa with normal morphology in samples frozen in LCCY was 60.4 &plusmn, 1.72%, and with Steridyl, 72.4 &plusmn, 2.10% (p &lt, 0.01). Semen frozen in Steridyl showed good stimulation of respiration by 2.4-DNP, which indicates that oxidative phosphorylation was retained after freezing&ndash, thawing. No differences among the extenders were seen with the DNA integrity of spermatozoa. Six out of ten (60%) mares were pregnant after artificial insemination (AI) by LCCY frozen semen, and 9/12 (75%) by Steridyl frozen semen. No differences among extenders were seen in pregnancy rate. In conclusion, Steridyl was proven to be a good diluent for freezing stallion semen, even though it was developed for ruminants.

The aim of the study was to search for new mutations in the previously studied gene loci of follicle-stimulating hormone receptor (FSHR), inhibin α (INHA), inhibin β A (INHAB), prolactin (PRL), transition protein 2 (TNP2), and sperm flagella 2 (SPEF2) by sequencing, as well as the search for associations of previously identified mutations at these loci with fresh semen quality in Russian Holstein bulls. Phenotypic data from 189 bulls was collected. Data was analyzed for most bulls for three years of semen collection. The maximum value of each semen quality indicator (doublet ejaculate volume, sperm concentration, progressive motility and total number of spermatozoa) were selected. SNPs were identified in the FSHR, INHA, INHAB, TNP2, SPEF2 genes. The PRL gene did not have polymorphism. Significant (p &lt, 0.05) associations of polymorphisms in the FSHR gene with double ejaculate volume, concentration and total number of spermatozoa were identified. Polymorphism in the INHA gene was significantly associated (p &lt, 0.05) with sperm concentration. Polymorphism in the INHAB gene was significantly associated (p &lt, 0.05) with doublet ejaculate volume and total number of spermatozoa. Polymorphisms in the TNP2 and SPEF2 genes did not have significant associations with semen quality. The SNPs studied in our pilot work may be considered as candidate genetic markers in the selection of bulls.

The formation and functioning of the animal reproductive system occurs as a result of the coordinated interaction of a wide range of genes. The hormone inhibin (INHA) is a marker of Sertoli cell function and exocrine testicular function (spermatogenesis state). The hormone prolactin (PRL) is a regulatory factor in the male reproductive system and affects gametogenesis. The aim of the work was to search for associations of inhibin alpha (INHA) and prolactin (PRL) gens polymorphism in the population of Russian Holstein bulls. Sperm traits (doublet ejaculate volume, concentration, progressive motility) were evaluated over three years of bull use and the highest rates for each trait were selected. 32 bulls had contrasting traits. DNA was isolated from semen samples. Samples were sequenced using the Applied Biosystems 3500 Genetic Analyzer using the BigDye® Terminator v3.1 Sequencing Standard Kit according to the protocol. Genotyping was then performed by PCR-RFLP. Data was analyzed using ANOVA. All pairwise multiple comparison between means were conducted by t-test. No polymorphism was detected in two regions of the PRL gene. The frequency of genotype AA in INHA gene was 0.813, the genotype AG -0.187. The GG genotype has not been identified. Bulls with the genotype AG had a higher concentration of cells (1.64 ± 0.093 billion/ml) than bulls with the genotype AA (1.35 ± 0.058) (P˂0.05). No significant differences were found in the volume of ejaculate and motility. But bulls with the AG genotype had greater ejaculate volume and motility than bulls with the AA genotype. The study of polymorphism in the genes that determine the formation and functioning of the reproductive system will help to understand the basics of these processes, as well as use them as molecular genetic markers for culling young bulls according to sperm quality. Project № АААА-А18-118021990006-9.