MAJOR APPROACHES FOR GENERATING AND ANALYZING TRANSGENIC MICE - AN OVERVIEW
MAJOR APPROACHES FOR GENERATING AND ANALYZING TRANSGENIC MICE - AN OVERVIEW
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DOI：
10.1161/01.hyp.22.4.599
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发表时间：
1993-10-01
期刊：
影响因子：
8.3
通讯作者：
SIGMUND, CD
中科院分区：
文献类型：
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作者：
SIGMUND, CD
在过去的十年中，整个动物的基因转移技术的发展为研究人员提供了前所未有的机会，以探测体内复杂的调控系统。我们对基因表达和调节机制的理解取得了重大进展，人类疾病动物模型的开发也取得了重大进展，这只是这一技术如何影响医学科学的两个例子。转基因动物被定义为这样的动物，其中一段DNA已经物理整合到所有细胞的基因组中，包括生殖系，因此它可以作为简单的孟德尔性状传递给后代。DNA片段通常由完整的克隆基因、cDNA或通过重组DNA方法修饰的新基因组成。基因的全基因组克隆通常用于研究组织和细胞特异性表达和调节，或者可用于过表达基因产物。或者，一个基因的编码区可以用于另一个基因的转录调控区，使其在新的组织和细胞类型谱中表达。许多方法可用于引入DNA片段，包括直接显微注射单细胞受精胚胎、逆转录病毒介导的转移或胚胎干细胞中的基因转移。最常用于产生转基因动物和进行“基因添加”实验的技术是直接显微注射。或者，通过特异性靶向基因组中的整合位点在胚胎干细胞中进行基因转移来进行基因缺失或“敲除"。这种方法有可能用于产生特定的基因突变，并具有很大的适用性，为创造动物模型，模仿人类疾病（如囊性纤维化）。概述了可用于产生转基因动物的技术，不同的基因转移方法的相对优点，并简要描述了一些问题，可以解决使用转基因动物进行了讨论。
Over the past decade, the development of gene-transfer technology in whole animals has afforded unprecedented opportunities for investigators to probe complex regulatory systems in vivo. Important advances in our understanding of the mechanisms of gene expression and regulation and the development of animal models of human diseases are but two examples of how this technology has affected medical science. Transgenic animals are defined as animals in which a segment of DNA has been physically integrated into the genome of all cells, including the germ line, so that it can be transmitted to offspring as a simple Mendelian trait. The DNA segment generally consists of a whole cloned gene, cDNA, or a novel gene modified by recombinant DNA methodologies. Whole genomic clones of genes are often used to study tissue- and cell-specific expression and regulation or can be used to overexpress a gene product. Alternatively, the coding region of one gene can be used to the transcriptional regulatory region of another gene, causing it to be expressed in a new spectrum of tissues and cell types. A number of methods can be used to introduce the DNA segment, including direct microinjection of one-cell fertilized embryos, retroviral-mediated transfer, or gene transfer in embryonic stem cells. The technique most often used to generate transgenic animals and perform ''gene addition'' experiments is direct microinjection. Alternatively, gene deletions or ''knockouts'' are performed by gene transfer in embryonic stem cells by specifically targeting the site of integration in the genome. This methodology has the potential to be used to generate specific gene mutations and has great applicability for the creation of animal models that emulate human diseases (such as cystic fibrosis). An overview of the technology available for generating transgenic animals, the relative merits of the different gene-transfer methods, and a brief description of some of the questions that can be addressed using transgenic animals are discussed.