| This study was designed to get & examine the developmental ability of reconstituted embryos after inter-species somatic cell nuclear transfer. By serial researchs, including isolating and culturing in auricular dermal fibroblasts of rhesus monkey, investigating their morphological characterization in advance, and evaluating the efficiency of the oocyte recovery rates as well as oocyte quality after repeated superovulation and surgical recovery in rabbits, finally, we obtained reconstituted embryos, which between rhesus monkey's fibroblasts and rabbit's enucleated oocytes. The main contents and results were as following:1. Establishment of dermal fibroblast cell line of rhesus monkey for the donor of nuclear transfer1) Ear skin samples were obtained by biopsy from 6 rethsus monkeys (ranging from 6 months to 1 year old). The average survival rate of skin tissue fragments is 91.67%.2) Fibroblasts were harvested by using conventional procedure of tissue culture. DMEM , TCM199, RPMI1640, DMEM+F12 as basic mediums containing 10%FBS were selected to the primary culture. The survival rates of the skin tissue pieces in the four medium were 80%, 53%, 93%, 13% respectively. Theresults indicated that DMEM and RPMI1640 had the better effect under the same condition. The attachment and the proliferation of cells in vitro were rapid when cells cultured in RPMI1640 medium.3) The proliferation of cells was rapid when density of cell layer was in 2xl04-lxl05/cm2.4) When RPMI1640 supplemented with 10%, 15%, 20% FBS respectively was used to culture the cells, the counted number shows that different concentrations of serum have not significant difference on fibroblasts growth, but when cell cultured in supplemented with 0.5%serum medium, cells sustained survival, and the proliferation of cell were slowly.5) Cultured skin fibroblasts of rhesus monkey were preserved at liquid nitrogen with cryogen of 10%DMSO in RPMI1640 medium. The seeding efficiencies were not significant change when cells were thawed after Imth, 3mth, 6mth, 12mth (pX).05).6) Fibroblasts keep relatively stable diploid (2n=42) when cultured and generated before 10 generations, the percentage of normal diploid at 3,5,7,10 passages were 92%,89%,85.5%,81% respectively.7) cells were analyzed by immunohistoche mistry, and showed negtive for cytokeratin and postive for vimentin and type III collagen.2. Recovery of mature oocytes in rabbit for recipient of nuclear transferRabbits were superovulated by conventional six injections (3d) of follicle-stimulating hormone (FSH) or by a single subcutaneous injection of FSH dissolved in a 25% solution of polyvinylpyrrolidone (PVP), and non-superovulated females were control group. The oocytes were recovered from both superovulated and nontreated females. Rabbits were given the same gonadotrophin treatment and laparotomy up to three times at intervals of at least 50 days. Results indicate as follow:1) The two superovulatory groups compared with the nontreated group, the overall and available numbers of recovered oocytes were more than 3 times higher (p=0.001), and with no significant differences in two superovulatory groups (p=0.001).2) Among the living rabbits after 3 consecutive times laparotomy, the efficiency of recovery in FSH treatment group and control protocols did not differ (P > 0.05).However, at the FSH-PVP treatment group, the mean numbers in the third laparotomy was significantly lower than the first one (P <0.01).3-Nuclear transplantation and developmental ability of reconstituted embryosSkin fibroblast cells from rhesus monkey were used as sources of donor nuclei. Enucleated oocytes recovered from repeated superovulatory and laparotomy were used as recipient oocytes for nuclear transfer. This study was designed to orgnize the reconstituted embryos after inter-species somatic cell nuclear transfer technology, and to examine the potential developmental ability under various factors in vitro. Results are as follow:1) the cleavage rate of embryos with fibr...
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