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Asian-Australas J Anim Sci > Volume 21(5); 2008 > Article
Review Paper
Asian-Australasian Journal of Animal Sciences 2008;21(5): 745-753.
DOI: https://doi.org/10.5713/ajas.2008.r.04    Published online May 6, 2008.
Recent Progress in Biotechnology-based Gene Manipulating Systems to Produce Knock-In/Out Mouse Models
Woon Kyu Lee*, Joong Jean Park, Seok Ho Cha, Cheol-heui Yun
Correspondence:  Woon Kyu Lee,
Gene-manipulated mice were discovered for the first time about a quarter century ago. Since then, numerous sophisticated technologies have been developed and applied to answer key questions about the fundamental roles of the genes of interest. Functional genomics can be characterized into gain-of-function and loss-of-function, which are called transgenic and knock-out studies, respectively. To make transgenic mice, the most widely used technique is the microinjection of transgene-containing vectors into the embryonic pronucleus. However, there are critical drawbacks: namely position effects, integration of unknown copies of a foreign gene, and instability of the foreign DNA within the host genome. To overcome these problems, the ROSA26 locus was used for the knock-in site of a transgene. Usage of this locus is discussed for the gain of function study as well as for several brilliant approaches such as conditional/inducible transgenic system, reproducible/inducible knockdown system, specific cell ablation by Cre-mediated expression of DTA, Cre-ERTM mice as a useful tool for temporal gene regulation, MORE mice as a germ line delete and site specific recombinase system. Techniques to make null mutant mice include complicated steps: vector design and construction, colony selection of embryonic stem (ES) cells, production of chimera mice, confirmation of germ line transmission, and so forth. It is tedious and labor intensive work and difficult to approach. Thus, it is not readily accessible by most researchers. In order to overcome such limitations, technical breakthroughs such as reporter knock-in and gene knock-out system, production of homozygous mutant ES cells from a single targeting vector, and production of mutant mice from tetraploid embryos are developed. With these upcoming progresses, it is important to consider how we could develop these systems further and expand to other animal models such as pigs and monkeys that have more physiological similarities to humans.
Keywords: Transgenic; Knock In/Out; ROSA26; Tetraploid; ES Cells; Gene Manipulation
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