Transgenic mouse models

Interest in manipulating the genetic make-up of organisms has a long-standing history, beginning with selective breeding to the present cloning of mammals. The route to transgenesis can be traced to the convergence of gene transfer methods developed in mammalian cell culture systems and methods to manipulate early mouse embryos (see Refs. 1-3 for historical perspectives). The past two decades have witnessed three major technological breakthroughs that have significantly enhanced our abilities to alter the genetic make-up of mammals. First, in the early 1980s, it was shown that foreign DNA could be introduced into the mouse germline by directly injecting into pronuclei of one-cell mouse embryo [4]. This process of transforming the mouse genome by pronuclear injections was referred to as “transgenic” [5]. Transgenic technology, first demonstrated in the mouse, is now practiced widely in flies, worms, birds, fish, frogs, and mammals including mice, cows, sheep, and pigs. Second, in the late 1980s, it was shown that genes could be disrupted by a homologous recombination-based strategy in embryonic stem (ES) cells [6,7]. These ES cells are then used to generate mice that carry specific alterations in the germline [8,9]. This process of mutating genes was referred to as “gene knockout’'. Gene knockout technol-36 Shashikant and Ruddleogy has resulted in the creation of a large number of mutant mouse strains, which has provided valuable insights into gene function at the organismic level (reviewed in Ref. 10). Third, in the mid 1990s, it was shown that animals could be generated by replacing the zygotic nucleus with the somatic nucleus [11,12]. This process, referred to as cloning or nuclear transfer, was first demonstrated in sheep and later in other organisms including mice [13]. The cloning of mammals has provoked an intense debate on the technological and ethical implications for the human cloning. In addition to these advances, a number of molecular biological techniques have emerged, raising the possibility of more sophisticated genome modifications. Many of these manipulations are carried out extensively in mice. In this chapter, we review emerging technologies that will enable us to generate better mouse models for the study of gene function.