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Efficient generation of isogenic primary human myeloid cells using CRISPR-Cas9 ribonucleoproteins.
Hiatt, Joseph; Cavero, Devin A; McGregor, Michael J; Zheng, Weihao; Budzik, Jonathan M; Roth, Theodore L; Haas, Kelsey M; Wu, David; Rathore, Ujjwal; Meyer-Franke, Anke; Bouzidi, Mohamed S; Shifrut, Eric; Lee, Youjin; Kumar, Vigneshwari Easwar; Dang, Eric V; Gordon, David E; Wojcechowskyj, Jason A; Hultquist, Judd F; Fontaine, Krystal A; Pillai, Satish K; Cox, Jeffery S; Ernst, Joel D; Krogan, Nevan J; Marson, Alexander.
Affiliation
  • Hiatt J; Medical Scientist Training Program, University of California, San Francisco, San Francisco, CA 94143, USA; Biomedical Sciences Graduate Program, University of California, San Francisco, San Francisco, CA 94143, USA; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Microbiol
  • Cavero DA; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94143, USA; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
  • McGregor MJ; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Quantitative Biosciences Institute, QBI, University of California, San Francisco, San Francisco, CA 94158, USA.
  • Zheng W; Department of Medicine, Division of Experimental Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
  • Budzik JM; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
  • Roth TL; Medical Scientist Training Program, University of California, San Francisco, San Francisco, CA 94143, USA; Biomedical Sciences Graduate Program, University of California, San Francisco, San Francisco, CA 94143, USA; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Microbiol
  • Haas KM; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Quantitative Biosciences Institute, QBI, University of California, San Francisco, San Francisco, CA 94158, USA.
  • Wu D; Medical Scientist Training Program, University of California, San Francisco, San Francisco, CA 94143, USA; Biomedical Sciences Graduate Program, University of California, San Francisco, San Francisco, CA 94143, USA.
  • Rathore U; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94143, USA; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Meyer-Franke A; J. David Gladstone Institutes, San Francisco, CA 94158, USA.
  • Bouzidi MS; Vitalant Research Institute, San Francisco, CA 94118, USA; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
  • Shifrut E; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94143, USA; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Lee Y; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94143, USA; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Kumar VE; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94143, USA; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Dang EV; Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94143, USA.
  • Gordon DE; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Quantitative Biosciences Institute, QBI, University of California, San Francisco, San Francisco, CA 94158, USA.
  • Wojcechowskyj JA; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Quantitative Biosciences Institute, QBI, University of California, San Francisco, San Francisco, CA 94158, USA.
  • Hultquist JF; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Quantitative Biosciences Institute, QBI, University of California, San Francisco, San Francisco, CA 94158, USA; Divis
  • Fontaine KA; J. David Gladstone Institutes, San Francisco, CA 94158, USA.
  • Pillai SK; Vitalant Research Institute, San Francisco, CA 94118, USA; Department of Laboratory Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
  • Cox JS; Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA.
  • Ernst JD; Department of Medicine, Division of Experimental Medicine, University of California, San Francisco, San Francisco, CA 94143, USA.
  • Krogan NJ; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Cellular and Molecular Pharmacology, University of California, San Francisco, San Francisco, CA 94158, USA; Quantitative Biosciences Institute, QBI, University of California, San Francisco, San Francisco, CA 94158, USA. Elect
  • Marson A; J. David Gladstone Institutes, San Francisco, CA 94158, USA; Department of Microbiology and Immunology, University of California, San Francisco, San Francisco, CA 94143, USA; Innovative Genomics Institute, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Medicine, Universit
Cell Rep ; 35(6): 109105, 2021 05 11.
Article in En | MEDLINE | ID: mdl-33979618
ABSTRACT
Genome engineering of primary human cells with CRISPR-Cas9 has revolutionized experimental and therapeutic approaches to cell biology, but human myeloid-lineage cells have remained largely genetically intractable. We present a method for the delivery of CRISPR-Cas9 ribonucleoprotein (RNP) complexes by nucleofection directly into CD14+ human monocytes purified from peripheral blood, leading to high rates of precise gene knockout. These cells can be efficiently differentiated into monocyte-derived macrophages or dendritic cells. This process yields genetically edited cells that retain transcript and protein markers of myeloid differentiation and phagocytic function. Genetic ablation of the restriction factor SAMHD1 increased HIV-1 infection >50-fold, demonstrating the power of this system for genotype-phenotype interrogation. This fast, flexible, and scalable platform can be used for genetic studies of human myeloid cells in immune signaling, inflammation, cancer immunology, host-pathogen interactions, and beyond, and could facilitate the development of myeloid cellular therapies.
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Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Ribonucleoproteins / Genome / Myeloid Cells / CRISPR-Cas Systems Limits: Animals / Humans Language: En Journal: Cell Rep Year: 2021 Document type: Article
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Full text: 1 Collection: 01-internacional Database: MEDLINE Main subject: Ribonucleoproteins / Genome / Myeloid Cells / CRISPR-Cas Systems Limits: Animals / Humans Language: En Journal: Cell Rep Year: 2021 Document type: Article