Method for promoting cow in-vitro embryo culture germ cell development
A technology of embryo culture medium and culture medium, which is applied to promote the in vitro maturation of oocytes, agricultural-animal husbandry and veterinary reproduction, in vitro fertilization and embryo culture, and the application of related culture medium in the cultivation of bovine in vitro fertilized embryos to promote the development of fertilized eggs, In the field of bovine in vitro fertilized embryo culture, it can solve the problems of low efficiency, high production cost of in vitro fertilized embryos, and restrictions on wide application
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Embodiment 1
[0053] Embodiment 1: the culture method of bovine in vitro fertilization embryo (no selenium in vitro)
[0054] 1. Reagents
[0055] In the specific test of the present invention, if not otherwise specified, the relevant reagents used are described in detail as follows:
[0056] Add double-antibody normal saline: normal saline containing 400 IU / mL penicillin and 400 μg / mL streptomycin.
[0057] Egg washing solution: BY basal culture solution supplemented with 3mg / mL bovine serum albumin.
[0058] Maturation medium: BY basal medium supplemented with 100 mL / L FBS, 10 μg / mL FSH, 10 μg / mL LH, 1 μg / mL E2, and 20 ng / mL EGF.
[0059] HEPES maturation medium: BY basal medium supplemented with 15mmol / L HEPES, 100mL / L FBS, 10μg / mL FSH, 10μg / mL LLH, 1μg / mL E2, and 20ng / mL EGF.
[0060] Fertilization medium: containing 112.0mM sodium chloride, 4.02mM potassium chloride, 2.25mM calcium chloride dihydrate, 0.52mM magnesium chloride hexahydrate, 0.83mM potassium dihydrogen phosphate, 37...
Embodiment 2
[0093] Embodiment 2: the culture method of bovine in vitro fertilization embryo (living body without selenium)
[0094] 1. Reagents
[0095] In the specific test of the present invention, if not otherwise specified, the relevant reagents used are described in detail as follows:
[0096] Add double anti-physiological saline, egg washing solution: the same as in Example 1.
[0097] Maturation medium: BY basal medium supplemented with 100 mL / L FBS, 10 μg / mL FSH, 10 μg / mL LH, 1 μg / mL E2, and 20 ng / mL EGF.
[0098] HEPES maturation medium: BY basal medium supplemented with 15mmol / L HEPES, 100mL / L FBS, 10μg / mL FSH, 10μg / mL LLH, 1μg / mL E2, and 20ng / mL EGF.
[0099] Fertilization culture fluid, semen preparation culture fluid, embryo culture fluid, BY basal culture fluid: the same as in Example 1.
[0100] 2. Cattle in vitro fertilization and embryo culture:
[0101] Step (1), collection and in vitro maturation of oocytes
[0102] Live bovine eggs were collected, and the obtain...
Embodiment 3
[0129] Embodiment 3: the culture method of bovine in vitro fertilization embryo (isolated selenium copper)
[0130] The main difference between this example and the above example 1 is that 0.25 mg / L sodium selenite and 0.075 mg / L anhydrous copper sulfate are additionally added to the BY basal culture solution.
[0131] 1. Reagents
[0132] In the specific test of the present invention, if not otherwise specified, the relevant reagents used are described in detail as follows:
[0133] Add double anti-physiological saline, egg washing solution: the same as in Example 1.
[0134] Maturation medium: BY basal medium supplemented with 100 mL / L FBS, 10 μg / mL FSH, 10 μg / mL LH, 1 μg / mL E2, and 20 ng / mL EGF.
[0135] HEPES maturation medium: BY basal medium supplemented with 15mmol / L HEPES, 100mL / L FBS, 10μg / mL FSH, 10μg / mL LLH, 1μg / mL E2, and 20ng / mL EGF.
[0136] Fertilization culture fluid, semen preparation culture fluid, embryo culture fluid: the same as in Example 1.
[0137...
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