iKA-CRISPR hESCs for inducible and multiplex orthogonal gene knockout and activation
iKA-CRISPR hESCs for inducible and multiplex orthogonal gene knockout and activation
复制标题
用于诱导型和多重正交基因敲除和激活的 iKA-CRISPR hESC
DOI：
10.1002/1873-3468.13127
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发表时间：
2018-07-01
期刊：
影响因子：
3.5
通讯作者：
Rong, Zhili
中科院分区：
文献类型：
--
作者：
Ma, Shufeng;Lv, Jie;Rong, Zhili
人类胚胎干细胞（hESCs）在早期人类胚胎发育模拟、疾病建模和细胞治疗中有着广泛的应用。为了实现这些应用，我们建立了用于诱导和多重正交基因敲除和激活的hESCs，我们将其命名为iKA-CRISPR hESCs。在细胞中，当多西环素诱导的Cas9-p300蛋白与含有14-bp靶序列（14-bp gRNA）的短向导RNA或含有20-bp长的gRNA结合时，分别可以激活基因转录或切割基因组DNA。我们还利用iKA-CRISPR技术证明，敲除OCT4促进了hESCs的分化，并且可以有效地激活与发育相关的microrna和转录因子。因此，iKA-CRISPR hESCs为控制基因表达网络提供了便利的平台，从而促进了hESCs在基础和转化生物医学研究中的应用。
Human embryonic stem cells (hESCs) have a wide range of applications in early human embryonic development mimics, disease modeling, and cell therapy. To fulfill these applications, we established hESCs for inducible and multiplex orthogonal gene knockout and activation, which we named iKA-CRISPR hESCs. In cells, when complexed with a short guide RNA containing a 14-bp target sequence (14-bp gRNA) or a long 20-bp gRNA, the doxycycline-induced Cas9-p300 protein could activate gene transcription or cleave genomic DNA, respectively. We also demonstrate using iKA-CRISPR hESCs that knockout of OCT4 promoted differentiation, and developmentally relevant microRNAs and transcription factors could be efficiently activated. Thus, iKA-CRISPR hESCs provide a convenient platform to control gene expression networks and, therefore, facilitate the applications of hESCs in basic and translational biomedical research.