Another way in which gene function can be analyzed is by adding DNA sequences of interest to the genome of an organism that normally lacks such sequences and then observing the effect of the introduced sequence on the organism’s phenotype. This method is a form of reverse genetics. An organism that has been permanently altered by the addition of a DNA sequence to its genome is said to be transgenic, and the foreign DNA that it carries is called a transgene (Figure 14.21). Here, we consider techniques for the creation of transgenic mice, which are often used in the study of the function of human genes because they can be genetically manipulated in ways that are impossible with humans and, as mammals, they are more similar to humans than are fruit flies, fish, and other model genetic organisms.

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The oocytes of mice and other mammals are large enough that DNA can be injected into them directly. Immediately after penetration by a sperm, a fertilized mouse egg contains two pronuclei, one from the sperm and one from the egg; these pronuclei later fuse to form the nucleus of the embryo. Mechanical devices can manipulate extremely fine, hollow glass needles to inject DNA directly into one of the pronuclei of a fertilized egg (Figure 14.22). Typically, a few hundred copies of cloned linear DNA are injected into a pronucleus. In a few of the injected eggs, copies of the cloned DNA integrate randomly into one of the chromosomes through a process called nonhomologous recombination. After injection, the embryos are implanted in a pseudopregnant female: a surrogate mother that has been physiologically prepared for pregnancy by mating with a vasectomized male.

Only about 10% to 30% of the injected embryos survive, and of those that do survive, only a few have a copy of the cloned DNA stably integrated into a chromosome. Nevertheless, if several hundred embryos are injected and implanted, there is a good chance that one or more mice whose chromosomes contain the foreign DNA will be born. Moreover, because the DNA was injected at the one-cell stage of the embryo, these mice usually carry the cloned DNA in every cell of their bodies, including their reproductive cells, and will therefore pass the foreign DNA on to their progeny. Through interbreeding, a strain of mice that carry the foreign gene can be created.

Transgenic mice have proved useful in the study of gene function. For example, proof that the SRY gene (see Chapter 4) is the male-determining gene in mice was obtained by injecting a copy of the SRY gene into XX embryos and observing that these mice developed as males. In addition, researchers have created a number of transgenic mouse strains that serve as experimental models for human genetic diseases.