College of Animal Science and Technology, Hebei Agricultural University, Baoding, PR China.
College of Life Science and Technology, Southwest Minzu University, Chengdu, China.
Anim Biotechnol. 2020 Apr;31(2):155-163. doi: 10.1080/10495398.2018.1559846. Epub 2019 Feb 8.
Somatic cell nuclear transfer (SCNT) technology has been applied in the construction of disease model, production of transgenic animals, therapeutic cloning, and other fields. However, the cloning efficiency remains limited. In our study, to improve SCNT efficiency, brilliant cresyl blue (BCB) staining were chosen to select recipient oocytes. In addition, DNA methyltransferase inhibitor Zebularine (5 nmol/L) and histone deacetylase inhibitor Scriptaid (0.2 μmol/L) were jointly used to treat sheep donor cumulus cells and reconstructed embryo. Moreover, the expression levels of embryonic development-related genes ( and ) of reconstructed embryo were also detected. Using BCB + oocytes as recipient cell, donor cumulus cells and reconstructed embryos were treated with 5 nmol/L Zebularine and 0.2 μmol/L Scriptaid, the blastocyst rate in Zeb + SCR-SCNT group (28.25%) was significantly higher than SCNT (21.16%) ( < 0.05). Furthermore, results showed that expression levels of and genes in Zeb + SCR-SCNT embryos were more similar to IVF embryos. Our study proved that 5 nmol/L Zebularine and 0.2 μmol/L Scriptaid treating with sheep donor cumulus cells and reconstructed embryos improved SCNT blastocyst rate and relieve the abnormal expression of embryonic developmental related genes.
体细胞核移植 (SCNT) 技术已应用于疾病模型构建、转基因动物生产、治疗性克隆等领域。然而,克隆效率仍然有限。在我们的研究中,为了提高 SCNT 效率,选择使用亮甲酚蓝 (BCB) 染色来选择受体卵母细胞。此外,还共同使用 DNA 甲基转移酶抑制剂 zebularine (5 nmol/L) 和组蛋白去乙酰化酶抑制剂 Scriptaid (0.2 μmol/L) 处理绵羊供体卵丘细胞和重构胚胎。此外,还检测了重构胚胎的胚胎发育相关基因 (和) 的表达水平。使用 BCB + 卵母细胞作为受体细胞,用 5 nmol/L zebularine 和 0.2 μmol/L Scriptaid 处理供体卵丘细胞和重构胚胎,Zeb + SCR-SCNT 组的囊胚率 (28.25%) 明显高于 SCNT 组 (21.16%) ( < 0.05)。此外,结果表明,Zeb + SCR-SCNT 胚胎中 和 基因的表达水平与 IVF 胚胎更为相似。本研究证明,用 5 nmol/L zebularine 和 0.2 μmol/L Scriptaid 处理绵羊供体卵丘细胞和重构胚胎可提高 SCNT 囊胚率,并缓解胚胎发育相关基因的异常表达。