Gene Editing in Human Pluripotent Stem Cells: Choosing the Correct Path
Gene Editing in Human Pluripotent Stem Cells: Choosing the Correct Path
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人类多能干细胞的基因编辑：选择正确的路径
DOI：
10.15436/2471-0598.15.004
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发表时间：
2015
期刊：
影响因子：
--
通讯作者：
D. Allison
中科院分区：
文献类型：
--
作者：
Amar M. Singh;Valeriya V Adjan Steffey;Tseten Yeshi;D. Allison
最近靶向核酸酶的出现为利用人类多能干细胞(HPSCs)进行基因修饰开辟了新的机会。这些修改的范围从创建常规的基因敲除到更具挑战性的单点突变。对于新用户和现有用户来说，由于新的和改进的技术正在迅速和持续地发展，决定具体修改兴趣的最佳方法可能是一项艰巨的任务。试剂和方法学之间的选择完全取决于实验的最终目标和要修改的轨迹。研究人员需要从锌指核酸酶(ZFN)、转录激活子样效应子核酸酶(TALEN)和规则间隔短回文重复序列(CRISPR)/Cas9中选择最好的核酸酶用于每个实验，这样可以在最少陷阱的情况下获得最高的成功可能性。此外，与第一代核酸酶相比有了重大改进，例如开发了二聚体CRISPR RNA引导的Fok1核酸酶(RFN，市场名称为NextGen™CRISPR)，它降低了“脱靶”突变率，为研究人员提供了更多的选择。如果研究人员需要进行点突变，那么必须考虑使用单链寡核苷酸(SSODN)作为同源定向修复的供体，还是使用供体载体中的选择盒与仅切除的猪Bac™转座酶相结合来留下无缝编辑。在这篇综述中，我们将提供当前技术的一般概述，以及产生点突变的方法，同时考虑它们的优缺点。
The recent emergence of targeted nucleases has opened up new opportunities for performing genetic modifications with human pluripotent stem cells (hPSCs). These modifications can range from the creation of a routine knock-out to the more challenging single point-mutation. For both the new and established user, deciding on the best approach for the specific modification of interest can be an arduous task, as new and improved technologies are rapidly and continuously being developed. The choices between the reagents and methodologies depends entirely on the end-goal of the experiments and the locus to be modified. Investigators need to decide on the best nuclease to use for each experiment from among Zinc-Finger Nucleases (ZFNs), Transcription Activator-Like Effector Nucleases (TALENs) and Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR)/Cas9 that would result in the highest likelihood of success with the fewest pitfalls. Furthermore, there have been significant improvements over the first-generation nucleases, such as the development of the dimeric CRISPR RNA-guided Fok1 nucleases (RFNs, marketed as NextGEN™ CRISPR) that reduces the “off-target” mutation rate, providing further options for investigators. Should researchers need to perform a point mutation, then considerations must be made between using single-stranded oligo-deoxynucleotides (ssODN) as the donor for homology-directed repair or utilizing a selection cassette within a donor vector in combination with an excision-only piggyBac™ transposase to leave a seamless edit. In this review, we will provide a general overview of the current technologies, along with methodologies for generating point mutations, while considering both their pros and cons.
一个 iCRISPR 平台，用于在人类多能干细胞中进行快速、可多重、可诱导的基因组编辑。
影响因子：
23.9
作者：
Gonzalez, Federico;Zhu, Zengrong;Shi, Zhong-Dong;Lelli, Katherine;Verma, Nipun;Li, Qing V.;Huangfu, Danwei
通讯作者：
Huangfu, Danwei
影响因子：
8.8
作者：
Firth AL;Menon T;Parker GS;Qualls SJ;Lewis BM;Ke E;Dargitz CT;Wright R;Khanna A;Gage FH;Verma IM
通讯作者：
Verma IM
人类多能细胞中的信号网络串扰：SMAD2/3调节的开关，可控制自我更新和分化之间的平衡。
影响因子：
23.9
作者：
Singh AM;Reynolds D;Cliff T;Ohtsuka S;Mattheyses AL;Sun Y;Menendez L;Kulik M;Dalton S
通讯作者：
Dalton S