4.1 Article

Generation and immunofluorescent validation of gene knockouts in adult human colonic organoids using multi-guide RNA CRISPR-Cas9

期刊

STAR PROTOCOLS
卷 4, 期 1, 页码 -

出版社

ELSEVIER
DOI: 10.1016/j.xpro.2022.101978

关键词

-

向作者/读者索取更多资源

In this study, a multi-guide RNA CRISPR-Cas9 gene-editing protocol is described that enables efficient complete gene knockout in adult human colonic organoids. The protocol also details crucial steps for maximizing gene-editing efficacy through tissue harvesting and validating gene knockout using immunofluorescent staining of the organoids against the target protein.
While readily achieved in cell lines, the application of CRISPR-Cas9 gene editing in human-derived organoids suffers from limited efficacy and complex protocols. Here, we describe a multi-guide RNA CRISPR-Cas9 gene-editing protocol which efficiently achieves complete gene knockout in adult human colonic organoids. This protocol also describes crucial steps including how to harvest patient tissue to maximize gene-editing efficacy and a technique to validate gene knockout following editing with immunofluorescent staining of the organoids against the target protein.

作者

我是这篇论文的作者
点击您的名字以认领此论文并将其添加到您的个人资料中。

评论

主要评分

4.1
评分不足

次要评分

新颖性
-
重要性
-
科学严谨性
-
评价这篇论文

推荐

暂无数据
暂无数据