Ishibashi Shoko, Kroll Kristen L, Amaya Enrique
The Healing Foundation Centre, The Faculty of Life Sciences, University of Manchester, Manchester, England, UK.
Methods Mol Biol. 2012;917:185-203. doi: 10.1007/978-1-61779-992-1_11.
Here we present a protocol for generating transgenic embryos in Xenopus laevis and Xenopus tropicalis. The method includes three steps: (1) The preparation of high-speed egg extracts, which facilitates the replacement of protamines in sperm nuclei with nucleosomes and decondenses the chromatin of sperm nuclei; (2) The isolation of sperm nuclei; and (3) The mixing of sperm nuclei, restriction enzyme, and high-speed extract in vitro, following by nuclear transplantation into unfertilized eggs to generate the transgenic embryos. This procedure generates non-mosaic transgenic embryos at high frequency and efficiency.
在此，我们展示了一种在非洲爪蟾和热带爪蟾中生成转基因胚胎的方案。该方法包括三个步骤：（1）制备高速卵提取物，这有助于用核小体取代精子细胞核中的鱼精蛋白并使精子细胞核的染色质解聚；（2）分离精子细胞核；以及（3）在体外将精子细胞核、限制酶和高速提取物混合，随后将细胞核移植到未受精卵中以生成转基因胚胎。此程序能以高频率和高效率产生非嵌合转基因胚胎。
