Abstract
Completion of the Xenopus laevis genome sequence from inbred J strain animals has facilitated the generation of germline mutant X. laevis using targeted genome editing. In the last few years, numerous reports have demonstrated that TALENs are able to induce mutations in F0 Xenopus embryos, but none has demonstrated germline transmission of such mutations in X. laevis. In this report we used the oocyte host-transfer method to generate mutations in both tyrosinase homeologs and found highly-penetrant germline mutations; in contrast, embryonic injections yielded few germline mutations. We also compared the distribution of mutations in several F0 somatic tissues and germ cells and found that the majority of mutations in each tissue were different. These results establish that X. laevis J strain animals are very useful for generating germline mutations and that the oocyte host-transfer method is an efficient technique for generating mutations in both homeologs.
Highlights
Xenopus laevis has a long history as a vertebrate model system in biomedical research, and has made important contributions to many fields, including cell cycle, somatic cell nuclear transfer, embryonic development, and cell fate specification
We examined the efficiency of germline transmission of Transcription activator-like effector nucleases (TALENs)-induced mutations in F0 adult X. laevis J strain animals produced by oocyte host-transfer, and determined whether the same germ cell mutations were induced in somatic tissues
Recent sequencing of the X. laevis genome has shown that the homeologous tyrosinase genes are located on the 2S and 2L chromosomes (Xenbase)
Summary
Xenopus laevis has a long history as a vertebrate model system in biomedical research, and has made important contributions to many fields, including cell cycle, somatic cell nuclear transfer, embryonic development, and cell fate specification. Simple microinjection of TALEN mRNAs into X. laevis embryos is sufficient to induce site-specific lesions and can be designed to target either one or both alloalleles (Nakade et al, 2015; Sakane et al, 2013; Suzuki et al, 2013). These studies demonstrated that TALENs are capable of inducing mutations in both homeologous genes, injections into embryos were not sufficient to generate a null phenotype. This result probably reflects the time required for the TALEN mRNAs to be translated during the rapid cell divisions of early Xenopus development
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