Abstract
Frog species of the genus Xenopus are widely used for studies of cell and developmental biology, and recent genome sequencing has revealed interesting phylogenetic relationships. Here we describe methods to generate haploid, triploid, and hybrid species starting from eggs and sperm of Xenopus laevis and Xenopus tropicalis that enable investigation of how genome size and content affect physiology at the organismal, cellular, and subcellular levels.
Highlights
Central to evolution, the biological consequences of changes in ploidy or genome hybridization are poorly understood
The most commonly used species, Xenopus laevis, is an allotetraploid with 36 chromosomes that arose through interspecific hybridization of diploid X. tropicalis-like progenitors that diverged from a common ancestor ~48 million years ago [11]
Whereas the hybrid produced when X. laevis eggs are fertilized by X. tropicalis sperm is viable, the reverse hybrid dies prior to gastrulation [8, 9]
Summary
The biological consequences of changes in ploidy or genome hybridization are poorly understood. Whereas the hybrid produced when X. laevis eggs are fertilized by X. tropicalis sperm (le×ts) is viable, the reverse hybrid (te×ls) dies prior to gastrulation [8, 9] In this protocol, we describe simple and optimized methods to generate haploid, triploid, and hybrid embryos of the Xenopus species, X. laevis and X. tropicalis (Fig. 1). Sperm irradiation inactivates the paternal genome to generate haploids, while cold shock of eggs blocks polar body extrusion to generate triploid embryos These are the simplest methods to perturb Xenopus embryo ploidy. We describe optimized methods to generate the viable hybrid obtained from the fertilization of X. laevis eggs with X. tropicalis sperm (le×ts) as well as the inviable hybrid obtained from the fertilization of X. tropicalis eggs with X. laevis sperm (te×ls), with reproducible fertilization efficiencies This protocol provides methods to prepare Xenopus embryos with varying genome sizes and content
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