Efficient gene knockout in goats using CRISPR/Cas9 system

PLoS One. 2014 Sep 4;9(9):e106718. doi: 10.1371/journal.pone.0106718. eCollection 2014.

Abstract

The CRISPR/Cas9 system has been adapted as an efficient genome editing tool in laboratory animals such as mice, rats, zebrafish and pigs. Here, we report that CRISPR/Cas9 mediated approach can efficiently induce monoallelic and biallelic gene knockout in goat primary fibroblasts. Four genes were disrupted simultaneously in goat fibroblasts by CRISPR/Cas9-mediated genome editing. The single-gene knockout fibroblasts were successfully used for somatic cell nuclear transfer (SCNT) and resulted in live-born goats harboring biallelic mutations. The CRISPR/Cas9 system represents a highly effective and facile platform for targeted editing of large animal genomes, which can be broadly applied to both biomedical and agricultural applications.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Alleles
  • Animals
  • Base Sequence
  • CRISPR-Cas Systems*
  • Embryo Transfer
  • Fibroblasts / cytology
  • Fibroblasts / metabolism*
  • Gene Knockout Techniques*
  • Genome*
  • Goats / genetics*
  • Lactoglobulins / deficiency
  • Lactoglobulins / genetics
  • Molecular Sequence Data
  • Mutation
  • Myostatin / deficiency
  • Myostatin / genetics
  • Nuclear Pore Complex Proteins / deficiency
  • Nuclear Pore Complex Proteins / genetics
  • Nuclear Transfer Techniques
  • Polymorphism, Restriction Fragment Length
  • Primary Cell Culture
  • Prions / antagonists & inhibitors
  • Prions / genetics

Substances

  • Lactoglobulins
  • Myostatin
  • Nuclear Pore Complex Proteins
  • Prions

Grants and funding

This work was supported by the National Basic Research Program of China (973 Program 2010CB530200 to CC), the Bingtuan Twelve Five-Year Special Projects (2012BB051 to CC), the National Natural Science Foundation of China (NSFC 31201800 and 31260534 to SH), and Utah Multidisciplinary Arrhythmia Consortium and Utah Agricultural Experimental Station (project1100 to IP). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.