Biallelic genome modification in F(0) Xenopus tropicalis embryos using the CRISPR/Cas system

Genesis. 2013 Dec;51(12):827-34. doi: 10.1002/dvg.22719. Epub 2013 Nov 21.

Abstract

Gene inactivation is an important tool for correlation of phenotypic and genomic data, allowing researchers to infer normal gene function based on the phenotype when the gene is impaired. New and better approaches are needed to overcome the shortfalls of existing methods for any significant acceleration of scientific progress. We have adapted the CRISPR/Cas system for use in Xenopus tropicalis and report on the efficient creation of mutations in the gene encoding the enzyme tyrosinase, which is responsible for oculocutaneous albinism. Biallelic mutation of this gene was detected in the F0 generation, suggesting targeting efficiencies similar to that of TALENs. We also find that off-target mutagenesis seems to be negligible, and therefore, CRISPR/Cas may be a useful system for creating genome modifications in this important model organism.

Keywords: CRISPR/Cas; TALENs; gene knockouts; gene targeting; zinc finger nucleases.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Albinism / genetics
  • Alleles
  • Animals
  • Base Sequence
  • CRISPR-Cas Systems*
  • Clustered Regularly Interspaced Short Palindromic Repeats*
  • Embryo, Nonmammalian / metabolism
  • Female
  • Gene Dosage
  • Gene Knockout Techniques
  • Genome
  • INDEL Mutation
  • Monophenol Monooxygenase / genetics*
  • Monophenol Monooxygenase / metabolism
  • Phenotype
  • Xenopus / embryology*
  • Xenopus / genetics*
  • Xenopus Proteins / genetics*
  • Xenopus Proteins / metabolism

Substances

  • Xenopus Proteins
  • Monophenol Monooxygenase