title: Construction of a CRISPR-Cas9 System for Pig Genome Targeting.
abstract: A Cas9/sgRNA RNA-guided endonuclease expression system including a codon-optimized Streptococcus pyogenes A20 Cas9 recombinant protein expression vector and a spacer-guide chimeric RNA expression vector using the porcine U6 promoter was constructed for application in pigs. Only the Flag2-NLS1-Cas9-NLS2 recombinant protein in complex with sgRNA was translocated into the nucleus; the Flag2-NLS1-Cas9-NLS2 protein alone was excluded from the nucleus. Up to 13% of porcine PK1 cells targeted in vitro were observed, regardless of transfection efficiency.
authors.affiliation: a Division of Animal Technology, Laboratories of Animal Technology, Agricultural Technology Research Institute, Xiangshan District , Hsinchu City , Taiwan.
authors.forename: Yu-Hsiu
authors.initials: YH
authors.lastname: Su
authors.forename: Tai-Yun
authors.initials: TY
authors.lastname: Lin
authors.forename: Chung-Lin
authors.initials: CL
authors.lastname: Huang
authors.forename: Ching-Fu
authors.initials: CF
authors.lastname: Tu
authors.forename: Chin-Kai
authors.initials: CK
authors.lastname: Chuang
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D001483:Base Sequence; D000076987:CRISPR-Associated Protein 9; D064113:CRISPR-Cas Systems / Q000235:genetics*; D002460:Cell Line; D004720:Endonucleases / Q000235:genetics; D005818:Genetic Engineering / Q000379:methods*; D016678:Genome / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D008969:Molecular Sequence Data; D000094704:RNA, Guide, CRISPR-Cas Systems / Q000235:genetics; D034421:Sus scrofa / Q000235:genetics*; D013552:Swine; D014162:Transfection
chemical_list: D001426:Bacterial Proteins; D000094704:RNA, Guide, CRISPR-Cas Systems; D000076987:CRISPR-Associated Protein 9; C000606107:Cas9 endonuclease Streptococcus pyogenes; D004720:Endonucleases
keywords: CRISPR; Cas9; sgRNA