Bovine cloned embryo freezing solution, thawing solution, kit and bovine cloned embryo freezing and thawing methods

Abstract

The invention provides a bovine cloned embryo freezing solution, a bovine cloned embryo thawing solution, a kit and bovine cloned embryo freezing and thawing methods, and belongs to the in-vitro embryo freezing technology. The freezing method comprises the following steps: putting a bovine cloned embryo into a basic solution, and balancing the embryo at a temperature of 24-25 DEG C for 5-10 minutes; putting the processed embryo into a first vitrification freezing solution to balance the embryo at a temperature of 24-25 DEG C for 4-6 minutes, putting the processed embryo into a second vitrification freezing solution to balance the embryo at a temperature of 24-25 DEG C for 4-6 minutes and putting the processed embryo into a third vitrification freezing solution to balance the embryo at a temperature of 24-25 DEG C for 35-45 seconds, and finally putting the processed embryo into liquid nitrogen to freeze. And the frozen bovine cloned embryo is put in a first thawing solution, the embryois thawed at a temperature of 37-39 DEG C for 50-70s, the thawed embryo is put in a second thawing solution, the embryo is balanced at a temperature of 24-25 DEG C for 4-6 min, and finally placing theprocessed embryo is put in a base solution, and the embryo is balanced at a temperature of 24-25 DEG C for 4-6 min. With the method disclosed by the invention, the conception rate of the bovine cloned embryo is as high as 25%.

Description

technical field [0001] The invention belongs to in vitro embryo freezing technology, in particular to bovine cloned embryo freezing liquid, thawing liquid, kit and method for bovine cloned embryo freezing and thawing. Background technique [0002] Cryopreservation of human and animal embryos has become an essential part of the field of artificial reproduction. In the past two decades, the development of cryopreservation technology can also reflect the development level of artificial reproductive technology. Modern cryopreservation technology has been used to cryopreserve sperm, eggs and embryos at different stages. Vitrification, in particular, has not only been widely recognized by the medical community, but has gradually replaced the traditional slow freezing method and has become the best choice for freezing technology. Compared with the traditional slow freezing method, the vitrification process does not produce water crystals in the cell body, thereby avoiding damage t...

AI Technical Summary

Problems solved by technology

[0003] In vitro fertilized and cloned embryos are of poorer quality and less resistant to freezing compared to in vivo embryos

The low quality of embryos leads to a further decline in the resurrection rate and conception rate of embryos after freezing and thawing

The traditional slow freezing method cannot effectively cryopreserve this kind of embryo, which greatly limits the application of in vitro fertilization and cloning technology in livestock breeding

The inventor of the existing bovine embryo vitrification technology, G Vajta’s scheme is as follows: freezing liquid: 15% glycerol + 15% DMSO; 38°C; thawing liquid: 1M sucrose, 38°C; compared with the traditional slow freezing method Improve the freezing effect and conception rate of embryos in vivo, but the application effect on cloned embryos is not ideal, generally only 5% conception rate

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