Genetic tools and mutagenesis strategies based on transposable elements are currently under development with a vision to link primary DNA sequence information to gene functions in vertebrate models. By virtue of their inherent capacity to insert into DNA, transposons can be developed into powerful tools for chromosomal manipulations. Transposon-based forward mutagenesis screens have numerous advantages including high throughput, easy identification of mutated alleles, and providing insight into genetic networks and pathways based on phenotypes. For example, the Sleeping Beauty transposon has become highly instrumental to induce tumors in experimental animals in a tissue-specific manner with the aim of uncovering the genetic basis of diverse cancers. Here, we describe a battery of mutagenic cassettes that can be applied in conjunction with transposon vectors to mutagenize genes, and highlight versatile experimental strategies for the generation of engineered chromosomes for loss-of-function as well as gain-of-function mutagenesis for functional gene annotation in vertebrate models, including zebrafish, mice, and rats. Transposons are mobile genetic elements with the unique ability to change their position in the genome. Because transposon integration can occur at vast numbers of locations in the genome, transposition is inherently mutagenic. An entire array of sophisticated mutagenic and reporter cassettes has been established. Both loss-of-function and gain-of-function phenotypes can be induced by insertions of transposons carrying such cassettes. Transposon-based functional genomics approaches have uncovered genes and genetic pathways in many areas of physiology and pathophysiology, including embryonic development, drug sensitivity, and resistance and neurobiology. Transposon forward mutagenesis has mainly been done in mice, on a variety of cancer-predisposing genetic or environmental backgrounds, for many types of cancer, and for cancer drug-resistance mechanisms.
Bibliographical noteFunding Information:
K. Kawakami received support through grants 15H02370 and 16H01651 from MEXT and NBRP from Amed. D. A. Largaespada acknowledges support from the National Cancer Institute (R01-CA113636) and The American Cancer Society Research Professor Award (#123939).
© 2017 Elsevier Ltd
- animal models
- forward genetics
- genetic screens
- insertional mutagenesis
- stem cells