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Building a Better Gene Trap
Researchers improve technique for large-scale studies of gene function in mice
By Lone Frank

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In the monumental task of tracking down the biological functions of some 40,000 human genes, scientists have a new tool. An American research team has improved a technique for 'trapping' genes—a strategy for mutating large numbers of genes in mice and linking them to specific traits. The researchers demonstrated the potential value of the technique on signaling genes.

"The project began six years ago and grew out of an interest in finding new signaling pathways," says William Skarnes, of the University of California, Berkeley, who led the research. "Genes involved in signaling are typically cell surface receptors, which are important targets for disease treatment."

Skarnes and colleagues have accomplished three things: They improved the overall efficiency of the process, set up the first trap to capture only certain types of genes, and made it possible to trap genes that are expressed in very low levels.

"There is no doubt this is an important step forward for functional genomics," says Ian Jackson, of the Medical Research Council in Edinburgh. "It shows for the first time that gene-trapping can be used for large-scale applications and it will make things go faster." Jackson wrote a commentary that accompanies the report by Skarnes' group in Nature Genetics.

Gene-trapping techniques have been around in various forms for years. They are examples of reverse genetics: Researchers mutate individual genes in a model system and analyze the effects to learn about the function of the gene. A second approach is direct genetics. Here, researchers start with a mutant trait, or phenotype, and the abnormality becomes the basis for tracking down the mutated gene.

Genetic enhancement of the vestigial tail phenotype by introducing one mutant copy of the LRP6 gene trap insertion.

"Rivalling camps have promoted one or the other method," says Jackson. The new study, he adds, suggests the value of both methods: "The way to get the job done is not to compete but to let the various approaches complement each other." The physical framework of the gene trap is a collection of immature and undifferentiated mouse embryonic stem cells. The cells are exposed to a genetic construct—a vector—that inserts into DNA. When cells produce the proteins carried by the vector, researchers can detect a disruption in the active gene. They then isolate mutated cells and preserve them; or they inject the cells into early mouse embryos to engineer live mice whose phenotype can be studied in detail.

"Setting the trap for genes that initiate signaling pathways meant we had to single out genes that encode proteins destined to be secreted or inserted into the cell membrane," explains Kathy Pinson, who co-authored the study.

The researchers inserted 500 mutations in embryonic stem cells, hitting 43 novel genes. They created mouse strains from 60 mutations, some of which were scrutinized in the laboratory. Others were shipped to researchers who study the trapped genes.

Skarnes is building a library of frozen embryonic stem cell lines that can be turned into mice. He plans to make the library available to other researchers.
X-gal staining of E9.5d embryo showing reporter gene expression associated with a secretory trap insertion in a novel gene.

"These cell lines represent a tremendous resource," says Jackson. At present, about ten percent of all mouse genes have been mutated, but Skarnes intends to create mutations in all 40,000. The researchers have generated vectors that will selectively pick up other classes of genes, says Pinson.

Many studies—and the similarities between the mouse and human genomes—suggest that there is much to be learned from mice about the functions of human genes. "Ten years of functional mouse genetics have taught us that we can extrapolate directly from mice to humans," says Jackson. "The mouse is an excellent model for human physiological processes."

Skarnes points out that gene function can be studied using both genetic and biochemical approaches. With genetics, the traits resulting from a mutation indicate where in the body a gene is involved. But a biochemical approach is necessary to characterize the role of the gene product in a variety of cells.

Although his passion is genetics, Skarnes recognizes the need for both approaches. "Ultimately," he says, "the confluence of genetic and biochemical information will lead to biological insights and thereby to cures for disease."

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Mitchell, K.J. et al. Functional analysis of secreted and transmembrane proteins critical to mouse development. Nat Genet 28, 241-249 (July 2001).
Jackson, I.J. Mouse mutagenesis on target. Nat Genet 28, 198-200 (July 2001).

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