title: Efficient generation of gene-modified pigs via injection of zygote with Cas9/sgRNA.
abstract: Co-injection of zygotes with Cas9 mRNA and sgRNA has been proven to be an efficient gene-editing strategy for genome modification of different species. Genetic engineering in pigs holds a great promise in biomedical research. By co-injection of one-cell stage embryos with Cas9 mRNA and Npc1l1 sgRNA, we achieved precise Npc1l1 targeting in Chinese Bama miniature pigs at the efficiency as high as 100%. Meanwhile, we carefully analyzed the Npc1l1 sgRNA:Cas9-mediated on- and off-target mutations in various somatic tissues and ovaries, and demonstrated that injection of zygotes with Cas9 mRNA and sgRNA is an efficient and reliable approach for generation of gene-modified pigs.
authors: Department of Laboratory Animal Science, College of Basic Medical Sciences, Third Military Medical University, Chongqing 400038, China.
authors: Yong
authors: Y
authors: Wang
authors: MOE Key Laboratory of Model Animal for Disease Study, Model Animal Research Center of Nanjing University, National Resource Center for Mutant Mice, Nanjing 210061, China.
authors: Yinan
authors: Y
authors: Du
authors: State Key Laboratory of Reproductive Medicine, Department of Histology and Embryology, Nanjing Medical University, Nanjing 210029, China.
authors: Bin
authors: B
authors: Shen
authors: Department of Laboratory Animal Science, College of Basic Medical Sciences, Third Military Medical University, Chongqing 400038, China.
authors: Xiaoyang
authors: X
authors: Zhou
authors: Department of Immunology, College of Basic Medical Sciences, Third Military Medical University, Chongqing 400038, China.
authors: Jian
authors: J
authors: Li
authors: Department of Laboratory Animal Science, College of Basic Medical Sciences, Third Military Medical University, Chongqing 400038, China.
authors: Yu
authors: Y
authors: Liu
authors: State Key Laboratory of Reproductive Medicine, Department of Histology and Embryology, Nanjing Medical University, Nanjing 210029, China.
authors: Jianying
authors: J
authors: Wang
authors: MOE Key Laboratory of Model Animal for Disease Study, Model Animal Research Center of Nanjing University, National Resource Center for Mutant Mice, Nanjing 210061, China.
authors: Jiankui
authors: J
authors: Zhou
authors: MOE Key Laboratory of Model Animal for Disease Study, Model Animal Research Center of Nanjing University, National Resource Center for Mutant Mice, Nanjing 210061, China.
authors: Bian
authors: B
authors: Hu
authors: School of Laboratory Medicine, Wenzhou Medical University, Wenzhou, Zhejiang 325035, China.
authors: Nannan
authors: N
authors: Kang
authors: School of Laboratory Medicine, Wenzhou Medical University, Wenzhou, Zhejiang 325035, China.
authors: Jimin
authors: J
authors: Gao
authors: Department of Animal and Avian Sciences, University of Maryland, College Park, MD 27042, USA.
authors: Liqing
authors: L
authors: Yu
authors: 1] MOE Key Laboratory of Model Animal for Disease Study, Model Animal Research Center of Nanjing University, National Resource Center for Mutant Mice, Nanjing 210061, China [2] Shanghai Key Laboratory of Reproductive Medicine, Shanghai 200025, China.
authors: Xingxu
authors: X
authors: Huang
authors: Department of Laboratory Animal Science, College of Basic Medical Sciences, Third Military Medical University, Chongqing 400038, China.
authors: Hong
authors: H
authors: Wei
mesh_terms: D000483:Alleles; D000818:Animals; D030801:Animals, Genetically Modified; D001483:Base Sequence; D064113:CRISPR-Cas Systems / Q000235:genetics*; D005260:Female; D018390:Gene Targeting / Q000379:methods; D008565:Membrane Proteins / Q000737:chemistry / Q000235:genetics; D008845:Microinjections / Q000379:methods; D008969:Molecular Sequence Data; D009030:Mosaicism; D009928:Organ Specificity / Q000235:genetics; D010053:Ovary / Q000378:metabolism; D017393:RNA Editing; D012333:RNA, Messenger; D013552:Swine; D015053:Zygote / Q000378:metabolism*; D000094704:RNA, Guide, CRISPR-Cas Systems
chemical_list: D008565:Membrane Proteins; D012333:RNA, Messenger; D000094704:RNA, Guide, CRISPR-Cas Systems
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