快速建立人结直肠类器官敲除系。

Division of Regenerative Medicine & Ansary Stem Cell Institute, Department of Medicine, Weill Cornell Medicine, 1300 York Avenue, New York, NY 10065, USA.

Department of Internal Medicine, University of Michigan, 1500 E Medical Center Drive, Ann Arbor, MI 48109, USA.

STAR Protoc. 2022 Apr 12;3(2):101308. doi: 10.1016/j.xpro.2022.101308. eCollection 2022 Jun 17.

Human colonic organoids derived from biopsy or autopsy tissues are a vital tool to study mucosal homeostasis, model colonic diseases, and develop therapeutics. Rapid and reliable generation of knockout organoid lines from multiple donors enables analysis of specific gene functions. Here, we report protocols to produce colonic organoid knockout lines within 1 to 2 weeks using lentiviral delivery of CRISPR-Cas9, achieving knockout efficiency of 90% or greater. These lines are suitable for multi-lineage differentiation and downstream analysis. For complete details on the use and execution of this protocol, please refer to Gu et al. (2022).

从活检或尸检组织中获得的人类结直肠类器官是研究黏膜稳态、模拟结直肠疾病和开发治疗方法的重要工具。从多个供体中快速可靠地产生基因敲除类器官系，可用于分析特定基因的功能。在此，我们报告了使用慢病毒递送 CRISPR-Cas9 在 1 至 2 周内生成结直肠类器官基因敲除系的方案，实现了 90%或更高的基因敲除效率。这些系适合多谱系分化和下游分析。如需详细了解该方案的使用和执行，请参阅 Gu 等人（2022 年）的研究。