2.1. Stallions and Semen Collection
This study was performed using four fertile miniature
Caspian horses between 10 and 12 years of age from Animal
Breeding Center of Karaj, Iran. The stallions were fed
sufficient coastal Bermuda grass hay and grain mix to
maintain good body condition. Semen was collected from
these stallions on a regular basis (3 times a week) by using
a Missouri artificial vagina. Seven ejaculates per stallion
were processed. The four stallions’ semen was collected on
the same day (n ¼ 7 days of collection). After collection, raw
semen was filtered through gauze to exclude the gel fraction
of the ejaculate and immediately processed.