Yanbian Cattle Somatic Cell Cloning, Fusion And Activation Conditions

As the situation of international market competition is more and more impetuous, breeds of cattle in our country are based on military service or military-flesh service and military-milk service, so it is hard to hold a dominant role in international stage for our country cattle products. From the 21 century, the urgency of our country cattle breeds being upgraded is on the rise, then people pay much more important attention to a new high performance technique-somatic cell cloning technique.Yanbian Yellow Cattle is one of five local elite in our country, it possesses higher flesh quality, thus it has a great international market potential. Recent years the technology cost on protecting species and selection is much bigger. In order to accelerate the improving process and elevate its selection efficiency, we introduce somatic cell cloning technique. So that the job can successfully carry out, we should in advance research the key link of technique route in somatic cell cloning technique-fusion and activation, therefore, it can ensure the technique successfully apply to Yanbian Yellow Cattle protecting species and selection.In this research, we use fresh granulosa cells as donor cells; we use MⅡenucleated oocytes as recipient cells. We do the experiment by means of acupressure enucleated oocytes putting donor granulosa cells into yolk space. In this experiment, we can investigate suitable conditions of fusion and activation in Yanbian Yellow Cattle from six aspects-recovery time before fusion; electric field strength in fusion; concentration of mannitol; recovery time before activation; activation modality; activation time. They are shown that:(1) Recovery, time before fusion in 2h can remarkably improve the efficiency and quality of fusion, the fused rate and the abnormal rate is 80.8% and 16.6%, respectively;(2) Electric field strength in fusion with 1100V/cm, the development rate of 4-6cell and 8-16 cell is 61.9% and 32.3%, respectively, it is significantly higher than the other three groups(P＜0.05);(3) The concentration of mannitol with 0.25M, 0.27M, 0.30M in fused rate, cleavage rate, abnormal rate and the development rate of reconstructed embryos, there were no significant differences;(4) Recovery time before activation with 20min and 25min, cleavage rate is 67.6% and 71.6%, abnormal rate of reconstructed embryos is 33.6% and 29.3%, it is an ideal recovery time before activation in Yanbian Yellow cattle;(5) Activation modality with Ion+6-DMAP, the cleavage rate and the development rate of 8-16 cell of reconstructed embryos is no significantly different from the other activation modality, abnormal rate and the development rate of 2-4 cell is 26.6% and 60.7%, respectively;(6) Activation time with 4h, the development rate of 8-16 cell of reconstructed embryos is 39.8%, significantly higher than the 3h and 6h treatment groups (P＜0.05), it.is the best suitable activation time in somatic cell cloning in Yanbian Yellow cattle.We can conclude that recovery time before fusion in 2h, the concentration of mannitol with 0.30mol/L, electric field strength in fusion with 1100V/cm is the optional fusion conditions in somatic cell cloning in Yanbian Yellow eattle; recovery time before activation with 20min-25min, activation modality with Ion+6-DMAP, aetivation time with 4h is. an ideal condition in somatic cell cloning in Yanbian Yellow cattle.