Generation of mice from wild-type and targeted ES cells by nuclear cloning
Generation of mice from wild-type and targeted ES cells by nuclear cloning
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DOI：
10.1038/72753
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发表时间：
2000-02-01
期刊：
影响因子：
30.8
通讯作者：
Jaenisch, R
中科院分区：
文献类型：
--
作者：
Rideout, WM;Wakayama, T;Jaenisch, R
F1 v6.5 5-6 149 21（14%）0 4 4（19%）F1 v6. 5 sc 84 8-9a 46 6（13%）2 2 2（33%）F1 rtTA 2 ESTA SD-18 8-9a 32 7（22%）0 1 1（14%）总计227 34（15%）2 7（21%）7（21%）129 v 18.6 5-6 107 19（19%）1 4 0 129 J1 11 145 26（18%）2 4 0 129（高）J1 35 b 92 23（25%）10 0 0 129 LJG-13 8-9a 74 8（11%）4 0 0总计418 76（18%）17 8（11%）0（0%）（在含有1，000 U/ml白血病抑制因子（LIF），但仅含5%胎牛血清的标准ES培养基中培养1-5天，不含饲养细胞）转化为去核的B6 D2 F1卵母细胞5。我们通过暴露于含Sr++的培养基来激活卵母细胞，然后将其在体外培养3天，然后转移到2.5天后的coblasts（dpc）假孕瑞士女性。我们在18.5-19.5 dpc对所有受体雌性进行剖腹产，然后培养任何活的幼崽。F1为129 SvJae × C57 BL/6遗传背景。存活的克隆动物都来自供体F1 ES细胞核，因为它们都有无菌的毛色，而供体卵母细胞来自黑色的非无菌的B6 D2 F1雌性，克隆胚胎已被转移到假孕的白色瑞士雌性中。靶向的ES细胞系是斜体的。在4-5代时靶向或亚克隆aES细胞，并在移植前再生长4代。b高传代。c仅移植已达到桑椹胚或囊胚期的胚胎。
F1 v 6.5 5–6 149 21 (14%) 0 4 4 (19%) F1 v6. 5 sc84 8–9a 46 6 (13%) 2 2 2 (33%) F1 rtTA2∆ SD-18 8–9a 32 7 (22%) 0 1 1 (14%) Total 227 34 (15%) 2 7 (21%) 7 (21%)129 v 18.6 5–6 107 19 (19%) 1 4 0 129 J1 11 145 26 (18%) 2 4 0 129 (high) J1 35b 92 23 (25%) 10 0 0 129 LJG-13 8–9a 74 8 (11%) 4 0 0 Total 418 76 (18%) 17 8 (11%) 0 (0%)We produced embryos by transfer of nuclei from ES cells (cultured for 1–5 d without feeder cells in standard ES media containing 1,000 U/ml leukaemia inhibitory factor (LIF), but only 5% fetal calf serum) into enucleated B6D2F1 oocytes5. We activated the oocytes by exposure to Sr++-containing media, then cultured them in vitro for 3 d before transfer into 2.5-days post-coitum (dpc) pseudopregnant Swiss females. We performed caesarean sections at 18.5–19.5 dpc for all recipient females followed by fostering of any live pups. F1 refers to the 129SvJae× C57BL/6 genetic background. Surviving cloned animals were all derived from donor F1 ES-cell nuclei because all had agouti coat colour, whereas donor oocytes came from black, non-agouti B6D2F1 females and the cloned embryos had been transferred into pseudopregnant white Swiss females. Targeted ES-cell lines are italicized. aES cells were targeted or subcloned at 4–5 passages and grown for another 4 passages before transplantation. bHigh passage. cOnly embryos which had reached morula or blastocyst stage were transferred.