Special culture medium and method for culturing porcine trophoderm stem cells
A technology for culturing trophoblast stem cells and stem cells, which is applied in the field of stem cell technology research, can solve the problems of low culture efficiency and failure to prove the characteristics of stem cells, and achieve the effects of low cell apoptosis rate, long-term stable stem cell characteristics, and strong proliferation and passaging ability
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Embodiment 1
[0047] Example 1: Isolation, culture and identification of porcine trophoblast stem cells from blastocysts derived from artificial insemination (AI)
[0048] 1. Establishment of the culture system of porcine trophoblast stem cells derived from AI-derived blastocysts
[0049] 1.1 Obtaining method of blastocyst:
[0050] The common estrous sows were bred by AI method, and the blastocysts were collected by the method of in vitro punching after the bridging. The specific operation is: select sows with high gestational age to be slaughtered on the sixth or seventh day after mating, take out the uterus, fix the front end of the uterine horn with a hemostatic forceps, inject egg flushing solution (DPBS+PVA) from the umbrella of the oviduct, and use The catheter is collected at the uterine horn, filtered and cleaned with the egg washing solution, and the blastocysts are collected under a stereo microscope, and transported to the laboratory at a constant temperature of 39°C (artif...
Embodiment 2
[0078] Example 2: Isolation, culture and identification of porcine trophoblast stem cells derived from somatic cell nuclear transfer (SCNT) blastocysts
[0079] 1. Establishment of culture system for porcine trophoblast stem cells isolated from SCNT blastocysts
[0080] 1.1 Acquisition of SCNT blastocysts: using a micromanipulator to perform nuclear transfer of porcine fibroblasts to obtain cloned embryos, acquisition and maturation of porcine oocytes, cultivation of donor fibroblasts, enucleation and injection of oocytes, The fusion and activation of oocytes and somatic cells were performed according to existing literature standards (Li et al., 2013, PMID: 24033142). The embryo culture medium PZM-3 was used to culture the reconstituted embryos until the sixth or seventh day, and the cysts were collected. Embryo. Compared with AI-derived blastocysts, its volume is smaller, the total number of cells in blastocysts is less, and the quality of embryos is poor (SCNT blastocysts a...
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