Obtaining Xenopus laevis Embryos

Abstract

The embryos of the African clawed frog, Xenopus laevis, are a powerful substrate for the study of complex fundamental biological and disease mechanisms in neurobiology, physiology, molecular biology, cell biology, and developmental biology. A simple and straightforward technique for generating a large number of developmentally synchronized embryos is in vitro fertilization (IVF). IVF permits simultaneous fertilization of thousands of eggs but requires the death of the parental male, which may not be feasible if the male comes from a stock of precious animals. An alternative to euthanizing a precious male is to use a natural mating, which allows for the collection of many embryos with minimal preparation but with the potential loss of the experimental advantage of developmental synchronization. Here we present both strategies for obtaining X. laevis embryos.

Deep learning is widely applicable to phenotyping embryonic development and disease

“…F0 founders were generated by injecting one-cell X. laevis J strain embryos (RRID: NXR 0024) with both T1 and T2 sgRNAs; 750 pg of each sgRNA and 1500 pg Cas9 were injected at 10 nl per embryo. Animals were housed in recirculating aquatic systems at the National Xenopus Resource (NXR) (RRID:SCR_013731); husbandry methods and parameters can be found elsewhere ( McNamara et al, 2018 ; Shaidani et al, 2020 , 2021a , b ). Fifteen F0 founders survived through metamorphosis; however, one was lost to bloating and apparent polycystic kidneys.…”

Cryopreservation of Xenopus Sperm and In Vitro Fertilization Using Frozen Sperm Samples

“…1. Euthanize the male frog(s) painlessly according to local regulations and remove testes as in Protocol: Obtaining Xenopus laevis Embryos (Shaidani et al 2021) or Protocol: Obtaining Xenopus tropicalis Embryos by In Vitro Fertilization . Use tweezers to carefully remove any blood vessels and excess fat tissue.…”

“…Extrude approximately 500 X. laevis or X. tropicalis eggs into a dry 90-mm Petri dish as described in Protocol: Obtaining Xenopus laevis Embryos (Shaidani et al 2021) or in Protocol: Obtaining Xenopus tropicalis Eggs . Check egg quality under a dissecting microscope as there is no point wasting frozen sperm on poor-quality eggs.…”

“…14. Culture the embryos as described in Protocol: Obtaining Xenopus laevis Embryos (Shaidani et al 2021) In addition, consider the following.…”

“…X. laevis males can be primed with 35-50 IU of PMSG 3-5 d before harvesting testes to improve sperm maturity, but this does not appear to improve X. tropicalis sperm. See Protocol: Obtaining Xenopus laevis Embryos(Shaidani et al 2021) for information on priming frogs. 100 Cite this protocol as Cold Spring Harb Protoc; doi:10.1101/pdb.prot107029 A. Noble et al…”