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Background  The RNA guided CRISPR/Cas9 nucleases have been proven to be effective for gene disruption in various animal models including  Xenopus tropicalis . The neural crest (NC) is a transient cell population during embryonic development and contributes to a large variety of tissues. Currently, loss-of-function studies on NC development in  X. tropicalis  are largely based on morpholino antisense oligonucleotide. It is worthwhile establishing targeted gene knockout  X. tropicails  line using CRISPR/Cas9 system to study NC development.    Methods  We utilized CRISPR/Cas9 to disrupt genes that are involved in NC formation in  X. tropicalis  embryos. A single sgRNA and  Cas9  mRNA synthesized in vitro, were co-injected into  X. tropicalis  embryos at one-cell stage to induce single gene disruption. We also induced duplex mutations, large segmental deletions and inversions in  X. tropicalis  by injecting  Cas9  and a pair of sgRNAs. The specificity of CRISPR/Cas9 was assessed in  X. tropicalis  embryos and the Cas9 nickase was used to reduce the off-target cleavages. Finally, we crossed the G0 mosaic frogs with targeted mutations to wild type frogs and obtained the germline transmission.    Results  Total 16 target sites in 15 genes were targeted by CRISPR/Cas9 and resulted in successful indel mutations at 14 loci with disruption efficiencies in a range from 9.3 to 57.8 %. Furthermore, we demonstrated the feasibility of generation of duplex mutations, large segmental deletions and inversions by using Cas9 and a pair of sgRNAs. We observed that CRISPR/Cas9 displays obvious off-target effects at some loci in  X. tropicalis  embryos. Such off-target cleavages was reduced by using the D10A Cas9 nickase. Finally, the Cas9 induced indel mutations were efficiently passed to G1 offspring.    Conclusion  Our study proved that CRISPR/Cas9 could mediate targeted gene mutation in  X. tropicalis  with high efficiency. This study expands the application of CRISPR/Cas9 platform in  X. tropicalis  and set a basis for studying NC development using genetic approach.    Electronic supplementary material  The online version of this article (doi:10.1186/s13578-016-0088-4) contains supplementary material, which is available to authorized users.

Efficient genome editing of genes involved in neural crest development using the CRISPR/Cas9 system in Xenopus embryos