title: Simultaneous generation of multi-gene knockouts in human cells.
abstract: Genome-editing techniques enable the generation of gene knockouts in various mammalian cell lines. However, it remains technically challenging to completely disrupt a targeted gene using a canonical method in a timely manner. To improve the efficiency of producing reliable genomic modifications, we designed a method using a linear donor fragment containing a reporter system. Combined with a homologous recombination-independent knock-in strategy, we successfully enriched those cell clones that specifically carry the target gene mutations. We observed a much improved success rate when generating single- and multiple-gene knockouts in a one-step procedure using this special protocol coupled with the CRISPR/Cas9 system. This new approach further empowers the molecular biological study of genes and their functions.
authors: Biodynamic Optical Imaging Center (BIOPIC), Beijing Advanced Innovation Center for Genomics, Peking-Tsinghua Center for Life Sciences, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing, China.
authors: Yuexin
authors: Y
authors: Zhou
authors: Biodynamic Optical Imaging Center (BIOPIC), Beijing Advanced Innovation Center for Genomics, Peking-Tsinghua Center for Life Sciences, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing, China.|Academy for Advanced Interdisciplinary Studies, Peking University, Beijing, China.
authors: Hongmin
authors: H
authors: Zhang
authors: Biodynamic Optical Imaging Center (BIOPIC), Beijing Advanced Innovation Center for Genomics, Peking-Tsinghua Center for Life Sciences, State Key Laboratory of Protein and Plant Gene Research, School of Life Sciences, Peking University, Beijing, China.
authors: Wensheng
authors: W
authors: Wei
mesh_terms: D000818:Animals; D001483:Base Sequence; D064113:CRISPR-Cas Systems / Q000235:genetics; D004260:DNA Repair / Q000235:genetics; D055786:Gene Knockout Techniques / Q000379:methods*; D057809:HEK293 Cells; D006367:HeLa Cells; D006801:Humans
keywords: 
  CRISPR
  ; knockout; nonhomologous end-joining