PARENT SESSION
Cryopreservation

DETECTION OF DNA DAMAGE IN RESPONSE TO COOLING INJURY IN EQUINE SPERMATOZOA USING SINGLE-CELL GEL ELECTROPHORESIS.

Linfor, Jennifer¹, Meyers, Stuart¹, ¹

ABSTRACT- Single cell gel electrophoresis (SCGE), or comet assay, has the ability to detect damage at the single cell level and has not been reported with use in equine sperm. The ability to detect nuclear damage at the single cell level could aid in the advancement of semen preservation. The goal of this experiment was to determine whether equine sperm undergo significant damage to DNA integrity following storage at decreased temperatures. Results revealed cells with fragmented tails of DNA, or comets, in the frozen-thawed treatments (1, 3, and 5-times) were significantly different from that of fresh sperm (p < 0.05). In addition, detected DNA damage was different for the three freeze-thaw treatments. Sensitivity of SCGE on equine sperm was further tested with ratios of known amounts of frozen-thawed and fresh cells. The amount of detectable DNA damage was positively correlated with the percentage of thawed sperm added to suspensions (r² = 0.92). Potential damage as a result of cooled storage was also investigated and SCGE revealed that damage increased linearly with increased time of storage. Detection of sublethal and/or uncompensible fertility factors in semen such as DNA fragmentation could be useful to detect differences in cryopreservation potential on an individual male, and add an important parameter to monitoring and preserving fertility on an individual basis.

KEY WORDS: stallion sperm, cryopreservation, DNA integrity