XB-ART-6130Dev Dyn. December 1, 2002; 225 (4): 422-33.
Xenopus, the next generation: X. tropicalis genetics and genomics.
A small, fast-breeding, diploid relative of the frog Xenopus laevis, Xenopus tropicalis, has recently been adopted for research in developmental genetics and functional genomics. X. tropicalis shares advantages of X. laevis as a classic embryologic system, but its simpler genome and shorter generation time make it more convenient for multigenerational genetic, genomic, and transgenic approaches. Its embryos closely resemble those of X. laevis, except for their smaller size, and assays and molecular probes developed in X. laevis can be readily adapted for use in X. tropicalis. Genomic manipulation techniques such as gynogenesis facilitate genetic screens, because they permit the identification of recessive phenotypes after only one generation. Stable transgenic lines can be used both as in vivo reporters to streamline a variety of embryologic and molecular assays, or to experimentally manipulate gene expression through the use of binary constructs such as the GAL4/UAS system. Several mutations have been identified in wild-caught animals and during the course of generating inbred lines. A variety of strategies are discussed for conducting and managing genetic screens, obtaining mutations in specific sequences, achieving homologous recombination, and in developing and taking advantage of the genomic resources for Xenopus tropicalis.
PubMed ID: 12454920
Article link: Dev Dyn.
Grant support: EY-06675 NEI NIH HHS , EY-10283 NEI NIH HHS , RR-13221 NCRR NIH HHS
Genes referenced: actc1 lgals4.2 pax6
Article Images: [+] show captions
|Figure 1. Adult Xenopus laevis and Xenopus tropicalis appear similar, except for size. Female X. laevis (left) and female X. tropicalis (right).|
|Figure 2. A comparison of embryonic development between X. laevis (A–D) and X. tropicalis (E–H). A,E: Vegetal view of early gastrula stage embryos, future dorsal is at the top. Arrowheads mark dorsal lip of the blastopore. The anterior of the embryo is toward the top in the next four panels. B,F: Dorsal view of neural plate stage embryos. The darkened stripe in the center of the embryo is the presumptive floorplate. C,G: Dorsal view of neural tube stage embryos. Arrows mark the region of neural-fold fusion. D,H: A lateral view of tadpole stage embryos; anterior is to the right. Magnification is the same at each stage.|
|Figure 3. Pax-6/green fluorescent protein (GFP) transgenics show complex patterns of gene expression in later development, demarcating numerous subdomains within the brain, spinal cord, and sensory tissues. A: A group of Pax-6/GFP transgenic F2 siblings at late swimming tadpole stages (Nieuwkoop and Faber stage 43–45; Nieuwkoop and Faber, 1956). Note the consistency of GFP pattern and intensity between tadpoles. B: A high-magnification view of the head of a Pax-6/GFP primary transgenic tadpole at stage 46. Pax-6/GFP is expressed in the brain (br), eye (ey; GFP fluorescence is blocked in this image by the pigmented retina), and olfactory placodes (olf). Arrows point to the olfactory nerve (between olfactory placodes and brain) and optic nerve (between eye and brain), which show low, but detectable, levels of GFP fluorescence. Small clusters of cells on the skin, which may be part of the lateral line (Winklbauer, 1989), also express GFP.|
|Figure 4. A triple transgenic X. tropicalis tadpole. Red fluorescence in the muscle is produced by expression of a transgene coupling the cardiac actin promoter with red fluorescent protein (RFP). Green fluorescence in the brain is produced by the Pax-6 promoter coupled to green fluorescent protein (GFP). The yellow color of the lens of the eye is produced by the combination of Pax-6/GFP expression in the neural retina and gamma-crystallin/RFP expression in the lens.|
|Figure 5. A putative X. tropicalis ear mutant uncovered by gynogenesis. A normal tadpole, stage 42, is on the left (A) and a homozygous mutant tadpole on the right (B). Note the enlarged otic vesicles (compare white outlines) and lack of otoconia (white calcifications within the inner ear, marked by black arrows) in the mutant.|