Calf in vitro embryo culture solution

A technology of embryo culture medium and culture medium, which is applied in the field of culture medium, can solve the problems of reducing the blastocyst development rate, and achieve the effect of high blastocyst development rate

Inactive Publication Date: 2011-02-02
北京奶牛中心
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, the addition of cysteamine should be appropriate,

Method used

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  • Calf in vitro embryo culture solution
  • Calf in vitro embryo culture solution

Examples

Experimental program
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Effect test

Embodiment 1

[0030] Example 1 Calf Hormone Superovulation and Egg Collection

[0031] On the day of calf superovulation, CIDR for sheep (for sheep, place of origin New Zealand) was implanted transvaginally with a suppository, and FSH (FSH, trade name Folltronpin-V, purchased from Canada Bioniche AnimalHealth Company), eggs were collected on the 7th day, and the thrombus was withdrawn at the same time.

[0032] The number of usable oocytes obtained in each calf super-rowing was 48 pieces.

Embodiment 2

[0033] Example 2 In vitro maturation of calf oocytes

[0034] 2 hours before the in vitro maturation operation, put an appropriate amount of maturation culture medium into each well of a 6-well conical culture plate (average 10 μl of maturation culture medium per oocyte), cover with mineral oil, and put it in a 38.5°C incubator for pre-equilibration . Among them, the composition of the mature medium is: TCM199+10mMHEPES+5%FBS+0.5μg / ml FSH+5μg / ml LH+1μg / ml E 2 + 27.5 μg / ml sodium pyruvate + 100 IU / ml penicillin + 100 μM cysteamine.

[0035] Put the identified and usable oocytes into the preheated maturation medium, wash them gently 4 times, transfer them into the maturation medium of a 6-well conical culture plate that has been equilibrated in the incubator for 2 hours, and place them in a constant temperature incubator Cultivate for 22h-24h. Culture conditions are 38.5°C, 5% CO 2 , saturated humidity. figure 1 Shown is the oocyte obtained after hormone superovulation of t...

Embodiment 3

[0036] Example 3 In vitro fertilization of calf oocytes

[0037] First, mature oocytes were mixed with fertilization fluid (114.0mM NaCl+4.02mM KCl+2.25mM CaCl 2 .2H 2 O+0.52mM MgCl 2 6H 2 O+0.83mM NaH 2 PO 4 h 2 O+37.0mM NaHCO 3 + 13.9mM glucose + 0.5mM sodium pyruvate + 3mg / mlBSA + 10μg / ml phenol red + 100IU / ml penicillin + 10μg / ml heparin), wash 2-3 times, and then put into the pre-balanced 2h fertilized fluid Medium (15 oocytes are placed in every 50ul fertilized fluid).

[0038]Take a thin tube (0.25ml) of frozen semen (the semen comes from the frozen semen of bulls from the Beijing Dairy Cow Center), put it in a water bath at 37°C for about 10 seconds, take it out until there are bubbles floating up, dry it with a facial tissue, and disinfect it with an alcohol cotton ball slim tube. The balanced centrifuge tube (containing semen washing solution: 114.0mM NaCl+4.02mM KCl+2.25mM CaCl 2 .2H 2 O+0.52mM MgCl 2 6H 2 O+0.83mM NaH 2 PO 4 h 2 O+37.0mM NaHCO 3 +1...

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Abstract

The invention provides a calf in vitro embryo culture solution containing cysteamine. The early-stage embryo in vitro development culture solution is used to culture the calf in vitro embryo and can obtain higher blastaea developmental rate.

Description

technical field [0001] The invention relates to a culture solution, in particular to a calf in vitro embryo culture solution. Background technique [0002] Like most mammals, bovine follicles occur in the fetal period, and the presence of antral follicles can be detected in the ovary of the fetus in the late pregnancy and a few days after birth, and the number reaches about 50 at the age of 2 months, which is obviously high. in adult cows. Therefore, making full use of the physiological characteristics that the production efficiency of calf oocytes is several times higher than that of adult cows, and exerting the maximum genetic potential of fine-bred calves has become one of the research hotspots in the field of dairy cow breeding technology. [0003] Using calves as donors can not only provide a large number of oocyte resources for in vitro production of embryos, but also meet the needs of scientific research and production such as oocyte freezing, embryo segmentation, cl...

Claims

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Application Information

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IPC IPC(8): C12N5/073
Inventor 朱玉林吕小青薛建华宣柏华李艳华周高举吴胜权赵鹏梁鸿斌麻柱
Owner 北京奶牛中心
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