Casein anti-freezing agent for cryopreservation of donkey semen and preparation method thereof
A technology for cryopreservation and casein, which is applied in the field of casein antifreeze and its preparation, can solve the problems of long reproductive cycle of female animals, low artificial insemination fertilization rate, complicated production and operation of frozen semen of donkeys, etc.
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Embodiment 1
[0067] Each 100mL of antifreeze contains 1.0g of casein, 2.0g of glucose, 1.0g of lactose, 3.0g of HEPESBUFFER, 2.0mL of glycerin, 2mL of egg yolk, 450,000 IU of penicillin, 4.5g of gentamicin, and 20mg of amphotericin B. The balance is HANK solution, weigh each component and set aside;
[0068] 2) Prepare the above-weighed casein, glucose, lactose, and HEPES BUFFER with HANK solution to form a base solution;
[0069] 3) Add the penicillin, gentamicin, and amphotericin B weighed in step 1) to the base liquid prepared above to make liquid I;
[0070]4) Adjust the pH of the liquid I to 7.1, then filter and sterilize it, and put it in a refrigerator at 3°C for later use;
[0071] 5) Take the liquid I before use, add the egg yolk weighed in step 1) to it, and prepare liquid II to obtain the casein antifreeze agent for cryopreservation of donkey semen.
Embodiment 2
[0073] Each 100mL of antifreeze contains 2.0g of casein, 2.5g of glucose, 1.5g of lactose, 5.0g of HEPESBUFFER, 2.5mL of glycerin, 3mL of egg yolk, 500,000 IU of penicillin, 5g of gentamicin, 25mg of amphotericin B, and the rest The amount is the HANK solution, weigh each component and set aside;
[0074] 2) Prepare the above-weighed casein, glucose, lactose, and HEPES BUFFER with HANK solution to form a base solution;
[0075] 3) Add the penicillin, gentamicin, and amphotericin B weighed in step 1) to the base liquid prepared above to make liquid I;
[0076] 4) Adjust the pH of the liquid I to 7.5, then filter and sterilize it, and put it in a refrigerator at 4°C for later use;
[0077] 5) Take the liquid I before use, add the egg yolk weighed in step 1) to it, and prepare liquid II to obtain the casein antifreeze agent for cryopreservation of donkey semen.
Embodiment 3
[0079] Each 100mL of antifreeze contains 3.0g of casein, 3.0g of glucose, 2.0g of lactose, 6.0g of HEPESBUFFER, 3.0mL of glycerin, 4mL of egg yolk, 550,000 IU of penicillin, 5.5g of gentamicin, and 30mg of amphotericin B. The balance is HANK solution, weigh each component and set aside;
[0080] 2) Prepare the above-weighed casein, glucose, lactose, and HEPES BUFFER with HANK solution to form a base solution;
[0081] 3) Add the penicillin, gentamicin, and amphotericin B weighed in step 1) to the base liquid prepared above to make liquid I;
[0082] 4) Adjust the pH of the liquid I to 7.8, then filter and sterilize it, and put it in a refrigerator at 5°C for later use;
[0083] 5) Take the liquid I before use, add the egg yolk weighed in step 1) to it, and prepare liquid II to obtain the casein antifreeze agent for cryopreservation of donkey semen.
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