Homology-mediated end joining-based targeted integration using CRISPR/Cas9
Yao, Xuan1,2; Wang, Xing1,2; Hu, Xinde1; Liu, Zhen1; Liu, Junlai2,3,4; Zhou, Haibo1; Shen, Xiaowen1; Wei, Yu1,5; Huang, Zijian1,2; Ying, Wenqin1; Wang, Yan1; Nie, Yan-Hong1; Zhang, Chen-Chen1; Li, Sanlan1; Cheng, Leping1; Wang, Qifang1; Wu, Yan6; Huang, Pengyu3; Sun, Qiang1; Shi, Linyu1; Yang, Hui1
Source PublicationCELL RESEARCH
AbstractTargeted integration of transgenes can be achieved by strategies based on homologous recombination (HR), microhomology-mediated end joining (MMEJ) or non-homologous end joining (NHEJ). The more generally used HR is inefficient for achieving gene integration in animal embryos and tissues, because it occurs only during cell division, although MMEJ and NHEJ can elevate the efficiency in some systems. Here we devise a homology-mediated end joining (HMEJ)-based strategy, using CRISPR/Cas9-mediated cleavage of both transgene donor vector that contains guide RNA target sites and similar to 800 bp of homology arms, and the targeted genome. We found no significant improvement of the targeting efficiency by the HMEJ-based method in either mouse embryonic stem cells or the neuroblastoma cell line, N2a, compared to the HR-based method. However, the HMEJ-based method yielded a higher knockin efficiency in HEK293T cells, primary astrocytes and neurons. More importantly, this approach achieved transgene integration in mouse and monkey embryos, as well as in hepatocytes and neurons in vivo, with an efficiency much greater than HR-, NHEJ- and MMEJ-based strategies. Thus, the HMEJ-based strategy may be useful for a variety of applications, including gene editing to generate animal models and for targeted gene therapies.
Keywordhomology-mediated end joining CRISPR/Cas9 monkey embryos neurons knock-in
Indexed BySCI ; CSCD
Funding ProjectShanghai City Committee of science and technology project[16JC1420202] ; Shanghai City Committee of science and technology project[14140900100]
WOS Research AreaCell Biology
WOS SubjectCell Biology
WOS IDWOS:000402545200010
Original Document TypeArticle
Citation statistics
Cited Times:138[WOS]   [WOS Record]     [Related Records in WOS]
Document Type期刊论文
Corresponding AuthorSun, Qiang; Shi, Linyu; Yang, Hui
Affiliation1.Chinese Acad Sci, Shanghai Inst Biol Sci, CAS Ctr Excellence Brain Sci & Intelligence Techn, Key Lab Primate Neurobiol,Inst Neurosci,State Key, Shanghai 200031, Peoples R China
2.Univ Chinese Acad Sci, Coll Life Sci, Beijing 100049, Peoples R China
3.ShanghaiTech Univ, Sch Life Sci & Technol, Shanghai 201210, Peoples R China
4.Chinese Acad Sci, Shanghai Inst Biol Sci, Inst Biochem & Cell Biol, Shanghai 200031, Peoples R China
5.Shanghai Univ, Shanghai 200444, Peoples R China
6.Natl Inst Biol Sci, Beijing 102206, Peoples R China
Recommended Citation
GB/T 7714
Yao, Xuan,Wang, Xing,Hu, Xinde,et al. Homology-mediated end joining-based targeted integration using CRISPR/Cas9[J]. CELL RESEARCH,2017,27(6):801-814.
APA Yao, Xuan.,Wang, Xing.,Hu, Xinde.,Liu, Zhen.,Liu, Junlai.,...&Yang, Hui.(2017).Homology-mediated end joining-based targeted integration using CRISPR/Cas9.CELL RESEARCH,27(6),801-814.
MLA Yao, Xuan,et al."Homology-mediated end joining-based targeted integration using CRISPR/Cas9".CELL RESEARCH 27.6(2017):801-814.
Files in This Item: Download All
File Name/Size DocType Version Access License
10.1038@cr.2017.76.p(7139KB)期刊论文出版稿开放获取CC BYView Download
Related Services
Usage statistics
Scholar Google
Similar articles in Scholar Google
[Yao, Xuan]'s Articles
[Wang, Xing]'s Articles
[Hu, Xinde]'s Articles
Baidu academic
Similar articles in Baidu academic
[Yao, Xuan]'s Articles
[Wang, Xing]'s Articles
[Hu, Xinde]'s Articles
Bing Scholar
Similar articles in Bing Scholar
[Yao, Xuan]'s Articles
[Wang, Xing]'s Articles
[Hu, Xinde]'s Articles
Terms of Use
No data!
Social Bookmark/Share
File name: 10.1038@cr.2017.76.pdf
Format: Adobe PDF
All comments (0)
No comment.

Items in the repository are protected by copyright, with all rights reserved, unless otherwise indicated.