Animal Science and Technology
Sartini, Becky, L
Fisheries, Animal, and Veterinary Science
epigenetics; DNA methylation; DNA; ram; spermatogenesis; gene expression
In the US livestock production industry, improving reproductive efficiency will improve animal welfare and maintain reasonable costs of meat and milk for consumers. In recent research, abnormalities in epigenetic markers in sperm during spermatogenesis, has been linked to male subfertility in many species. Epigenetics is the study of changes in organisms caused by modifications of gene expression, including DNA methylation, rather than alteration of the genetic code itself. When this process is disturbed, it can negatively impact semen therefore decreasing its fertility. Through further research on how DNA methylation influences gene expression during spermatogenesis and its impact on sperm quality, we can begin to build a comprehensive assay on sire fertility. In this study, the aim was to compare the methylation status of male germ cell genes in spermatozoa using a ram model. We hypothesized that there is variation in DNA methylation level of specific genes among individual rams. We chose to investigate the gene SIRT1, which is partially responsible for sperm maturation and capacitation, and methylation status, which has previously shown to be correlated with bull fertility. To date, a protocol to isolate genomic DNA in ram sperm after a Percoll wash to remove seminal plasma has been developed using the Qiagen DNA Blood and Tissue Kit (catalog number: 69504). From these experiments, we have routinely isolated an average of 20.38 ng/uL of DNA per sperm sample, with a range of 7.15 - 51.48 ng/uL for each collection. The average DNA purity number is 1.82, with a range of 1.5 - 2.63. Validation of the methylation specific polymerase chain reaction and comparison of amplification using primers that anneal to methylated and unmethylated DNA will determine if individual ram semen samples vary with DNA methylation.