Rapid and Cost-Effective Gene Targeting in Rat Embryonic Stem Cells by TALENs

被引：50

作者：

Tong, Chang ^{[1
]}

Huang, Guanyi ^{[1
]}

Ashton, Charles ^{[1
]}

Wu, Hongping ^{[1
]}

Yan, Hexin ^{[1
]}

Ying, Qi-Long ^{[1
]}

机构：

[1] Univ So Calif, Keck Sch Med, Dept Cell & Neurobiol, Eli & Edythe Broad Ctr Regenerat Med & Stem Cell, Los Angeles, CA 90033 USA

来源：

JOURNAL OF GENETICS AND GENOMICS | 2012年 / 39卷 / 06期

关键词：

TAL effector; BMPR2; Rat ES cells; Gene-targeting; KNOCKOUT RATS; HOMOLOGOUS RECOMBINATION; MICROINJECTION; MUTAGENESIS; NUCLEASES;

D O I：

10.1016/j.jgg.2012.04.004

中图分类号：

Q5 [生物化学]; Q7 [分子生物学];

学科分类号：

071010 ; 081704 ;

摘要：

The rat is the preferred animal model in many areas of biomedical research and drug development. Genetic manipulation in rats has lagged behind that in mice due to the lack of efficient gene targeting tools. Previously, we generated a knockout rat via conventional homologous recombination in rat embryonic stem (ES) cells. Here, we show that efficient gene targeting in rat ES cells can be achieved quickly through transcription activator-like effector nuclease (TALEN)-mediated DNA double-strand breaks. Using the Golden Gate cloning technique, we constructed a pair of TALEN targeting vectors for the gene of interest in 5 days. After gene transfection, the targeted rat ES cell colonies were isolated, screened, and confirmed by PCR without the need of drug selection. Our results suggest that TALEN-mediated gene targeting is a superior means of establishing genetically modified rat ES cell lines with high efficiency and short turnaround time.

引用

页码：275 / 280

页数：6

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