A simple and rapid approach to manipulate pseudorabies virus genome by CRISPR/Cas9 system
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  • 作者:Aotian Xu ; Chao Qin ; Yue Lang ; Mingyue Wang ; Mengyang Lin…
  • 关键词:CRISPR/Cas9 ; Genome engineering ; Knock in ; Knock out ; Large DNA virus ; Pseudorabies virus
  • 刊名:Biotechnology Letters
  • 出版年:2015
  • 出版时间:June 2015
  • 年:2015
  • 卷:37
  • 期:6
  • 页码:1265-1272
  • 全文大小:1,108 KB
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  • 作者单位:Aotian Xu (1)
    Chao Qin (1)
    Yue Lang (1)
    Mingyue Wang (1)
    Mengyang Lin (1)
    Chuang Li (1)
    Rui Zhang (1)
    Jun Tang (1)

    1. State Key Laboratory of Agrobiotechnology and College of Veterinary Medicine, China Agricultural University, Beijing, 100193, China
  • 刊物类别:Biomedical and Life Sciences
  • 刊物主题:Life Sciences
    Microbiology
    Biotechnology
    Applied Microbiology
    Biochemistry
  • 出版者:Springer Netherlands
  • ISSN:1573-6776
文摘
Objectives The broad host range of pseudorabies virus (PRV) and large capacity for foreign DNA make it a promising vector for the development of vaccines and agents of gene therapy. Results We show that up to 100?% viral gene disrupting efficiency was achieved by simple co-transfection of the purified PRV genomes with the clustered regularly-interspaced, short palindromic repeats (CRISPR)/CRISPR-associated protein 9 (Cas9) into cells. Furthermore, CRISPR/Cas9-mediated knock-in of >4-kb-long DNA cassettes into the PRV genome at a positive rate of 50?% by a homology-independent DNA repair mechanism without constructing homology arms. This approach requires only a simple plasmid construction and is applicable to knock-in of other foreign genes. Conclusion Our studies offered simple and efficient methods to manipulate PRV.

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