Department of Physiology, Catholic University of Daegu School of Medicine, Daegu, Korea.
Transgenic Res. 2010 Jun;19(3):437-47. doi: 10.1007/s11248-009-9327-3. Epub 2009 Oct 1.
A critical problem in the production of transgenic animals is the uncontrolled constitutive expression of the foreign gene, which occasionally results in serious physiological disorders in the transgenic animal. In this study, we report successful production of transgenic chickens that express the human erythropoietin (hEPO) gene under the control of a tetracycline-inducible promoter. A recombinant Moloney murine leukemia virus (MoMLV)-based retrovirus vector encapsidated with vesicular stomatitis virus G glycoprotein (VSV-G) was injected beneath the blastoderm of unincubated chicken embryos (stage X). Out of 198 injected eggs, 15 chicks hatched after 21 days of incubation and 14 hatched chicks expressed the vector-encoded hEPO gene when fed doxycycline, a tetracycline derivative, without any significant physiological dysfunctions. The expression of hEPO reverted to the pre-induction state by removing doxycycline from the diet. The biological activity of the hEPO produced in the transgenic chickens was comparable to commercially available CHO cell-derived hEPO. Successful germline transmission of the transgene was also confirmed in G1 transgenic chicks produced from crossing G0 transgenic roosters with non-transgenic hens. Tetracycline-inducible expression of the hEPO gene was also confirmed in the blood and eggs of the transgenic chickens.
生产转基因动物的一个关键问题是外源基因的非可控组成型表达，这偶尔会导致转基因动物出现严重的生理紊乱。在这项研究中，我们报告了成功生产出的转基因鸡，这些鸡在四环素诱导启动子的控制下表达人红细胞生成素（hEPO）基因。一种基于 Moloney 鼠白血病病毒（MoMLV）的逆转录病毒载体被包裹在水疱性口炎病毒 G 糖蛋白（VSV-G）中，然后注射到未孵化的鸡胚（X 期）的胚盘下。在 198 个注射的鸡蛋中，有 15 只小鸡在孵化 21 天后孵化出来，并且在喂食强力霉素（一种四环素衍生物）时，14 只孵化出来的小鸡表达了载体编码的 hEPO 基因，而没有任何明显的生理功能障碍。当从饮食中去除强力霉素时，hEPO 的表达恢复到诱导前的状态。在转基因鸡中产生的 hEPO 的生物活性与市售的 CHO 细胞衍生的 hEPO 相当。通过将 G0 转基因公鸡与非转基因母鸡杂交，也在 G1 转基因小鸡中证实了转基因的种系传递。在转基因鸡的血液和鸡蛋中也证实了 hEPO 基因的四环素诱导表达。
