Association of the polymorphism of the Pon-1 locus with the concentration of Paraoxonase-1 and post-thaw sperm viability in pig

Abstract

The objective of the present work was to correlate the expression of the PON-1 locus polymorphism, the concentration of the enzyme paraoxonase 1 in seminal plasma and the effect on post-thawing sperm viability in the pig. Nine ejaculates from 3 boars were used, to obtain seminal plasma; part of the ejaculates was centrifuged at 800 g/10 min; the material obtained was stored at -20°C until use. To freeze the semen, the samples were diluted 1:1 in commercial diluent (Magapor®) at 37°C and the temperature was gradually lowered to 16°C. They were transported to the laboratory protected from light, and kept at the same temperature in the laboratory for 24 h. After this, they were frozen using the glycerolation technique and a portion was cryopreserved by adding 20% of seminal plasma to the conservation medium and another portion was made without seminal plasma. The concentration of PON-1 was determined by an enzyme-linked immunosorbent assay (ELISA). The functional status of the plasma membrane was evaluated by chlortetracycline staining. The amplification of a fragment of the PON-1 gene was carried out by PCR and the identification of allelic variability by the analysis of Restriction Fragment Length Polymorphism (RFLP), with the enzymes HinfI and FokI, from a DNA sample of each individual. The results were analyzed by analysis of variance (ANOVA) and Spearman correlation, with the use of statistical software (Statistica V10 for Windows). In electrophoresis of enzymatic digestion with the enzyme HinfI All the expected fragments were obtained, with the FokI enzyme a mutation was observed in one individual. No correlation was observed (R=-0.1, P>0.05) between the concentration of the paraoxonase-1 enzyme in seminal plasma and the observed mutations. As well as the concentration of the enzyme paraoxonase-1 in the ejaculates was 1.8 to 2.7 ng/mL and this had no statistically significant difference in the seminal plasma of the ejaculates analyzed (P>0.05). As well as with sperm viability in thawed semen (P>0.05).