Association Between MTHFR Gene Polymorphism And Boar Semen Quality

Boar semen quality directly affects sow conception rate,litter size and progeny quality,which is essential for pig farm performance and economic efficiency.Studies have reported that 5,10-methylenetetrahydrofolate reductase?MTHFR?gene defects cause elevated levels of homocysteine?Hcy?,induce oxidative stress,and cause sperm obstruction and infertility are important enzymes in the metabolism of folic acid and Hcy,and play an important role in regulating intracellular folate levels,DNA synthesis and methylation.This paper aims to establish and detect Hcy content in boar semen and plasma,hydrogen peroxide?H₂O₂?and malondialdehyde?MDA?levels,and study single nucleotide polymorphisms of MTHFR gene?single?The relationship between nucleotide polymorphism,SNP and H₂O₂ levels and MDA levels in different types of boar seminal plasma,in order to screen the SNP locus of boar sperm MTHFR gene as a biomarker for early warning boar semen quality,for the boar station in advance The theoretical basis for the elimination of inferior boars.The main findings are as follows:1)Relationship between Hcy content of boar sperm and liquid quality:49 healthy Duroc,Landrace and Yorkshire were selected,and 49 boars were classified into semen quality according to recorded sperm density,effective sperm count and sperm motility?sperm?Viability>80%and sperm deformity rate<10%)and poor?sperm viability<70%or sperm deformity rate>20%?2 groups.Among them,33 boars with excellent semen quality and 16 poor pigs.The sperm density,effective sperm count and sperm motility of the semen with poor semen quality were significantly lower than those of the semen with excellent semen quality?P<0.01?,while the sperm deformity rate was significantly higher than that of the semen with excellent semen quality?P<0.01?..The Hcy levels of plasma and sperm were detected by high performance liquid chromatography and analyzed for their correlation with boar semen quality.The results showed that the Hcy content of spermatozoa in the group with poor semen quality was significantly higher than that in the group with excellent semen quality?P<0.01?,but there was no significant difference in plasma Hcy content?P>0.05?.2)Relationship between polymorphism of boar sperm MTHFR gene and semen quality:According to the record of the boar station,the semen quality of the boar with the semen utilization rate of 100%3 months before sampling is the best?mainly high sperm motility and 20 boars with low deformity rate,sperm motility>90%and sperm deformity rate<5%,and semen with less than 60%utilization rate have the worst semen quality?mainly low sperm motility and high deformity rate,sperm motility?20 boars<75%and sperm abnormality rate>15%?.Sperm DNA was extracted and amplified,and the MTHFR exon fragment was amplified.The PCR stocks of each of the 40 boar fragments were then mixed and the SNP locus on the MTHFR exon of the boar sperm was identified by a one-generation sequencing technique.The results showed that there were 13 SNPs in the exon of the MTHFR gene of boar sperm,two of which resulted in a change in the encoded amino acid;7 sites were consistent with the SNP site of the human MTHFR gene,no A similar site for MTHFR 677C>T associated with human infertility was found.Semen samples of 945 Duroc boars,459 heads of Landrace and 86 Yorkshire boars were collected,and the MTHFR gene SNP locus of 1490 boar sperm samples was detected by high-throughput SNP typing technique,and the amount of semen was measured.Correlation analysis was performed on indicators such as sperm density,total sperm count,effective sperm count,sperm motility,and sperm deformity rate.The results show:The locus c.450C>T in Duroc locus was significantly correlated with sperm motility?P<0.05?;c.294C>T locus was significantly correlated with total sperm count and effective sperm count?P<0.05?.The c.450C>T locus in Landrace boar was significantly correlated with semen volume;c.450C>T locus was significantly correlated with sperm density;c.1278G>A,c.1041G>A,c.570C>T and c.414C The>G locus was not only significantly associated with sperm density,but also significantly associated with sperm abnormality?P<0.05?.No SNP locus was found to be associated with semen quality in Yorkshire boars.6 of c.1485A>G,c.1287A>G,c.1278G>A,c.1041G>A,c.570C>T,and c.414C>G for the Landrace boar in accordance with Hada Weinberg's law The locus and the Yorkshire boar meet the Harder's law of Hardy.c.1485A>G,c.1287A>G,c.1278G>A,c.1041G>A,c.570C>T,c.450C>T,c The gene linkage disequilibrium analysis was performed at 7 sites of.414C>G.The results show:The six SNP loci on the MTHFR exon of Landrace boar sperm constitute two haplotype modules,and the c.1485A>G and c.1287A>G loci of module 1 are in strong linkage disequilibrium?D'>0.8,r2>0.33?;the four SNPs of c.1278G>A,c.1041G>A,c.570C>T,and c.414C>G of module 2 are in strong linkage disequilibrium?D'>0.8,r2>0.33?;7 SNPs on the MTHFR exon of the Yorkshire sperm constitute a haplotype block,c.1485A>G,c.1287A>G,c.1278G>A in the module The five SNP loci of c.1041G>A and c.570C>T are in strong linkage disequilibrium between the two?D'>0.8,r2>0.33?.3)A total of 170 Landrace boars were selected and the levels of H₂O₂ and MDA in seminal plasma were detected and correlated with the SNP locus of MTHFR gene.The results showed that:H₂O₂ levels and MDA in different genotypes of c.1278G>A,c.1041G>A,c.570C>T,c.414C>G and c.294C>T loci of Landrace boar MTHFR There is a significant difference in levels.In summary,the main conclusions of this study are as follows:the Hcy content of boar sperm in the poor quality semen group is significantly higher than that in the semen quality group,but there is no significant difference in plasma Hcy content.Landrace MTHFR c.1278G>A,c.1041G>A,c.570C>T,c.414C>G and c.294C>T loci different genotypes not only have significant differences with semen quality,but also with fine There is a significant difference in H₂O₂ levels and MDA levels in the pulp,so the polymorphisms at these loci may serve as potential biomarkers for predicting semen quality in Landrace boars.