title: Allele-specific genome editing and correction of disease-associated phenotypes in rats using the CRISPR-Cas platform.
abstract: The bacterial CRISPR/Cas system has proven to be an efficient gene-targeting tool in various organisms. Here we employ CRISPR/Cas for accurate and efficient genome editing in rats. The synthetic chimeric guide RNAs (gRNAs) discriminate a single-nucleotide polymorphism (SNP) difference in rat embryonic fibroblasts, allowing allele-specific genome editing of the dominant phenotype in (F344 × DA)F1 hybrid embryos. Interestingly, the targeted allele, initially assessed by the allele-specific gRNA, is repaired by an interallelic gene conversion between homologous chromosomes. Using single-stranded oligodeoxynucleotides, we recover three recessive phenotypes: the albino phenotype by SNP exchange; the non-agouti phenotype by integration of a 19-bp DNA fragment; and the hooded phenotype by eliminating a 7,098-bp insertional DNA fragment, evolutionary-derived from an endogenous retrovirus. Successful in vivo application of the CRISPR/Cas system confirms its importance as a genetic engineering tool for creating animal models of human diseases and its potential use in gene therapy.
authors: Institute of Laboratory Animals, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan.
authors: K
authors: K
authors: Yoshimi
authors: Institute of Laboratory Animals, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan.
authors: T
authors: T
authors: Kaneko
authors: Institute of Laboratory Animals, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan.
authors: B
authors: B
authors: Voigt
authors: Institute of Laboratory Animals, Graduate School of Medicine, Kyoto University, Kyoto 606-8501, Japan.
authors: T
authors: T
authors: Mashimo
mesh_terms: D054366:Agouti Signaling Protein / Q000235:genetics; D000483:Alleles; D000818:Animals; D064113:CRISPR-Cas Systems; D005818:Genetic Engineering / Q000379:methods*; D015316:Genetic Therapy / Q000379:methods; D014442:Monophenol Monooxygenase / Q000235:genetics; D010641:Phenotype; D020641:Polymorphism, Single Nucleotide; D019009:Proto-Oncogene Proteins c-kit / Q000235:genetics; D017393:RNA Editing; D000094704:RNA, Guide, CRISPR-Cas Systems; D051381:Rats
chemical_list: D054366:Agouti Signaling Protein; C515184:Asip protein, rat; D000094704:RNA, Guide, CRISPR-Cas Systems; D014442:Monophenol Monooxygenase; D019009:Proto-Oncogene Proteins c-kit