| Semen was collected from the Tibetan Mastiff and the method of semen collection; preservation extenders and semen cryopreservation were investigated to improve the quality of frozen sperm in the current research, aimed to identify the suitable method in the Tibetan mastiff semen cryopreservation.1. Semen cryopreservation using different extenders was researched, and the optimal extender was selected. The results showed that the extender No.2and the extender No.8were the best for diluting semen in the sperm viability, there was significantly different compared with control group(extender No.1) and other groups,while extender No.4,5and6were better than extender No.7,3and1; The extender No.2,8and6were the best for diluting semen in the intact acrosome rate, there was significantly different compared with control group(extender No.1) and other groups.At last, the extender No.2and extender No.8were the best.2. The methods of sperm collection were developed and semen evaluation was conducted59ejaculates (tried64times) were collected by the digital manipulation, as well as2ejaculates (tried20times) were obtained by an artificial vagina, the success rates were92.2%and10%, respectively. The Tibetan Mastiff aged2.5to5years, it is the best age of semen collection, there is a good breeding history. The semen characteristics in Tibetan Mastiff were evaluated, which include spermatozoa progressive motility(0.89～0.95),sperm valume(1.9～2.3ml), pH(6.5), sperm concentration(2.0～2.7×108个/ml), deformity rates (5.2～7.7%).3. In present study, some major technical aspect studies of frozen semen in Tibetan Mastiff were investigated which include different cooling time, different glycerol concentrations, different glycerol equilibration time, different distances from liquid nitrogen (LN) surface and different thawing temperatures, aimed to identify the suitable method in the Tibetan mastiff semen cryopreservation. The results showed that effects on freezing-thawing Tibetan Mastiff sperm was best when cooling time was200ml and250ml water, glycerol concentration was2%and3%, glycerol equilibration time was15min and30min, distance from liquid nitrogen (LN) surface was3cm and thawing temperature was40℃.The spermatozoa progressive motility was0.40using optimal solution after thawing. |
