The movement of mouse genetics from a backwater field of study to the forefront of modern biomedical research was catalyzed by the recombinant DNA revolution, which began 20 years ago and has been accelerating in pace ever since. With the ability to isolate cloned copies of genes and to compare DNA sequences from different organisms came the realization that mice and humans (as well as all other placental mammals) are even more similar genetically than they were thought to be previously. An astounding finding has been that all human genes have counterparts in the mouse genome which can almost always be recognized by cross-species hybridization. Thus, the cloning of a human gene leads directly to the cloning of a mouse homolog which can be used for genetic, molecular, and biochemical studies that can then be extrapolated back to an understanding of the function of the human gene. In only a subset of cases are mammalian genes conserved within the genomes of Drosophila or C. elegans.
This result should not be surprising in light of current estimates for the time of divergence of mice, flies and nematodes from the evolutionary line leading to humans. In general, three types of information have been used to build phylogenetic trees for distantly related members of the animal kingdom — paleontological data based on radiodated fossil remains, sequence comparisons of highly conserved proteins, and direct comparisons of the most highly conserved genomic sequences, namely the ribosomal genes. Unfortunately, flies (Drosophila) and nematodes (C. elegans) diverged apart from the line leading to mammals just prior to the time of the earliest fossil records in the Cambrian period which occurred 570 million years ago. The divergence of mice and people occurred relatively recently at 60 million years before present (see Section 2.2.1). These numbers are presented graphically in Figure 1.3, where a quick glance serves to drive home the fact that humans and mice are ten times more closely related to each other than either is to flies or nematodes.
Although the haploid chromosome number associated with different mammalian species varies tremendously, the haploid content of mammalian DNA remains constant at approximately three billion basepairs. It is not only the size of the genome that has remained constant among mammals; the underlying genomic organization (discussed in Chapter 5) has also remained the same as well. Large genomic segments — on average, 10-20 million basepairs — have been conserved virtually intact between mice, humans, and other mammals as well. In fact, the available data suggest that a rough replica of the human genome could be built by simply breaking the mouse genome into 130-170 pieces and pasting them back together again in a new order (Nadeau, 1984; Copeland et al., 1993). Although all mammals are remarkably similar in their overall body plan, there are some differences in the details of both development and metabolism, and occasionally these differences can prevent the extrapolation of mouse data to humans and vice versa (Erickson, 1989). Nevertheless, the mouse has proven itself over and over again as being the model experimental animal par excellence for studies of nearly all aspects of human genetics.
Among mammals, the mouse is ideally suited for genetic analysis. First, it is among the smallest mammals known with adult weights in the range of 25-40 g, 2,000-3,000-fold lighter than the average human adult. Second, it has a short generation time — on the order of 10 weeks from being born to giving birth. Third, females breed prolifically in the lab with an average of 5-10 pups per litter and an immediate postpartum estrus. Fourth, an often forgotten advantage is the fact that fathers do not harm their young, and thus breeding pairs can be maintained together after litters are born. Fifth, for developmental studies, the deposition of a vaginal plug allows an investigator to time all pregnancies without actually witnessing the act of copulation and, once again, without removing males from the breeding cage. Finally, most laboratory-bred strains are relatively docile and easy to handle.
The high resolution genetic studies to be discussed later in this book require the analysis of large numbers of offspring from each of the crosses under analysis. Thus, a critical quotient in choosing an organism can be expressed as the number of animals bred per square meter of animal facility space per year. For mice, this number can be as high as 3,000 pups/m2 including the actual space for racks (five shelves high) as well as the inter-rack space as well. All of the reasons listed here make the mouse an excellent species for genetic analysis and have helped to make it the major model for the study of human disease and normative biology.
The close correspondence discovered between the genomes of mice and humans would not, in and of itself, have been sufficient to drive workers into mouse genetics without the simultaneous development, during the last decade, of increasingly more sophisticated tools to study and manipulate the embryonic genome. Today, genetic material from any source (natural, synthetic or a combination of the two) can be injected directly into the nuclei of fertilized eggs; two or more cleavage-stage embryos can be teased apart into component cells and put back together again in new "chimeric" combinations; nuclei can be switched back and forth among different embryonic cytoplasma; embryonic cells can be placed into tissue culture, where targeted manipulation of individual genes can be accomplished before these cells are returned to the embryo proper. Genetically altered live animals can be obtained subsequent to all of these procedures, and these animals can transmit their altered genetic material to their offspring. The protocols involved in all of these manipulations of embryos and genomes have become well-established and cookbook manuals (Joyner, 1993; Wassarman and DePamphilis, 1993; Hogan et al., 1994) as well as a video guide to the protocols involved (Pedersen et al., 1993) have been published.
While it is likely that none of these manipulations has yet been applied to human embryos and genomes, it is ethical, rather than technical, roadblocks that impede progress in this direction. The mental image invoked is of a far more sophisticated technology than the so-called futuristic scenario of embryo farms described in Huxley's Brave New World (1932).
Progress has also been made at the level of molecular analysis within the developing embryo. With the polymerase chain reaction (PCR) protocol, DNA and RNA sequences from single cells can be characterized, and enhanced versions of the somewhat older techniques of in situ hybridization and immuno-staining allow investigators to follow the patterns of individual gene expression through the four dimensions of space and time (Wassarman and DePamphilis, 1993; Hogan et al., 1994). In addition, with the omnipresent micro-techniques developed across the field of biochemistry, the traditional requirement for large research animals like the rat, rabbit, or guinea pig has all but evaporated.
Finally, with the automation and simplification of molecular assays that has occurred over the last several years, it has become possible to determine chromosomal map positions to a very high degree of resolution. Genetic studies of this type are relying increasingly on extremely polymorphic microsatellite loci (Section 8.3) to produce anchored linkage maps (Chapter 9), and large insert cloning vectors — such as yeast artificial chromosomes (YACs) — to move from the observation of a phenotype, to a map of the loci that cause the phenotype, to clones of the loci themselves (Section 10.3). Thus, many of the advantages that were once uniquely available to investigators studying lower organisms, such as flies and worms, can now be applied to the mouse through the three-way marriage of genetics, molecular biology, and embryology represented in Figure 1.4 . It is the intention of this book to provide the conceptual framework and practical basis for the new mouse genetics.