Derivation of Germline Competent Rabbit Embryonic Stem Cell Lines

描述（由申请人提供）：由于其解剖学、遗传学和生物化学与人类相似，家兔是研究许多人类疾病的良好动物模型。它还用于药物筛选，抗体生产和治疗性蛋白质的生产。兔子足够大，可以为实验工作提供足够数量的组织，产生足够数量的抗体或治疗性蛋白质，但它又足够小，对大多数实验室来说是经济的。 目前，原核显微注射法仍是制备转基因兔的主要方法。没有可用的生殖系感受态兔胚胎干（rbES）细胞系。然而，原核显微注射效率低，导致随机整合，并且不能产生基因靶向的转基因动物。 本SBIR项目的目的是建立生殖系感受态rbES细胞系，用于生产基因靶向转基因兔。在第一阶段，我们建议制备兔胚胎成纤维细胞作为饲养层，合成重组兔白血病抑制（LIF）因子和衍生推定rbES细胞。我们假设，使用饲养层和LIF从兔源将增加获得生殖能力rbES细胞的机会。 我们预计在完成I期研究后，将获得至少20个推定的rbES细胞系。在第二阶段，我们将努力验证这些rbES细胞在囊胚注射后是否可以促进生殖系细胞，我们将努力从这些细胞中产生基因靶向转基因兔。本项目的成功将建立第一个非鼠源ES细胞系，提高兔的转基因效率，使基因靶向兔的生产成为可能。 兔是研究人类多种疾病的良好动物模型，因为它与人类在解剖学、遗传学和生物化学方面具有相似性。它还用于药物筛选，抗体生产和治疗性蛋白质的生产。兔尚未进入这些领域的主要原因是尚未建立具有生殖系能力的兔胚胎干细胞系。本项目的成功将建立第一个非鼠源ES细胞系，提高兔的转基因效率，使基因靶向兔的生产成为可能。

DESCRIPTION (provided by applicant): The rabbit is a good animal model for the study of many human diseases because of its anatomical, genetic and biochemical similarities to the human. It is also used for drug screening, production of antibody and production of therapeutic proteins. The rabbit is large enough to provide adequate quantities of tissue for experimental work, produce sufficient amount of antibody or therapeutic proteins, and yet it is small enough to be economical for most laboratories. To date, pronuclear microinjection remains the primary method to produce transgenic rabbits. There are no germline competent rabbit embryonic stem (rbES) cell lines available. Pronuclear microinjection, however, is of low efficiency, results in random integration, and cannot produce gene targeted transgenic animals. The purpose of this SBIR project is to establish germline competent rbES cell lines for the production of gene targeted transgenic rabbits. In Phase I we propose to prepare rabbit embryonic fibroblasts as feeder layers, synthesize recombinant rabbit leukemia inhibitory (LIF) factor and derive putative rbES cells. We hypothesize that using feeder layers and LIF from rabbit origin will increase the chance of getting germline competent rbES cells. We expect to derive at least 20 putative rbES cell lines upon completion of the Phase I study. In Phase II, we will work to validate if these rbES cells can contribute to germ line cells after blastocyst injection and we will work to produce gene targeted transgenic rabbits from these cells. The success of this project will establish the first germline competent ES cell lines in a non-murine species, improve the transgenic efficiency in rabbits, and make the production of gene targeted rabbits possible. The rabbit is a good animal model for the study of many human diseases because of its anatomical, genetic and biochemical similarities to the human. It is also used for drug screening, production of antibody and production of therapeutic proteins. The main reason that rabbit has not been in these fields is that there have been no established germline competent rabbit embryonic stem (rbES) cell lines. The success of this project will establish the first germline competent ES cell lines in a non-murine species, improve the transgenic efficiency in rabbits, and make the production of gene targeted rabbits possible.