如何高效制备生殖系传递的嵌合体小鼠

Efficiently Production of Chimeric Mice Competent for Germline Transmitted

目前,嵌合体技术已成为胚胎干细胞(ES细胞)和诱导多能性干细胞(iPS细胞)研究不可或缺的一部分,嵌合体小鼠模型主要用于基因功能、调控机制、致病机理等各方面的研究。要安全可靠地进行ES细胞和iPS细胞的各种应用性研究,首先需要验证细胞在体内向各种组织定向分化的能力,特别是生殖系传递的能力。而高效制备多能性干细胞生殖系传递的嵌合体小鼠,需要在分子水平、细胞水平、动物个体水平以及制备方法等各方面全面综合地分析考虑,才能达到实验设计的最优化。制备嵌合体小鼠主要有注射法和聚集法2种方法,各有利弊,在整个制备过程中,需要考虑不同发育阶段以及不同小鼠品系胚胎的分化潜能、选用远交系和杂交系的胚胎作为受体,同时要考虑ES细胞的遗传背景,选择低代次的远交系和杂交系的ES细胞或者iPS细胞,以及优选KSR培养体系。对于操作者,需根据实际情况,全方位地综合考虑,从而制定出合理的操作方案。

Chimeric mice capable of germline transmission are important tools for studying gene function, mechanisms of development, and disease pathogenesis. Chimeras are indispensable for research on embryonic stem cells （ESCs） and induced pluripotent stem cells （iPSCs）. When planning an experiment that includes production of chimeric mice, several factors should be considered that may have an impact on the molecular, cellular, or tissue/ organ levels. Recipient embryos at different stages of development have different efficiencies for donor cell engraftment and survival, leading to a wide range of levels of chimerism. Capability for tetraploid complementation is key, but so are other recipient embryo factors such as mitochondrial quantity and quality. Whether the recipient strain is inbred or out-breeding needs to also be considered. In the donor stem cells, genetic background is critically important, as are the cell culture conditions such as the KSR system used to engineer and expand donor cells before transplantation. The injection and aggregation methods are the two primary methods for producing chimeras, and both have advantages and disadvantages. This review is intended to assist investigators with a comprehensive assessment of what factors will be most important for meeting their specific goals in a chimeric mouse project.