Use of the heteroduplex mobility assay and cell sorting to select genome sequences of the CCR5 gene in HEK 293T cells edited by transcription activator-like effector nucleases

Genetics and Molecular Biology

Arildo Nerys-junior²  Lendel C. Costa²  Luciene P. Braga-dias¹  Márcia Oliveira¹  Átila D. Rossi²  Rodrigo Delvecchio Da Cunha²  Gabriel S. Gonçalves²  Amilcar Tanuri² 

[1] ,Universidade Federal do Rio de Janeiro Laboratório de Virologia Molecular Rio de Janeiro RJ ,Brazil

关键词: CCR5;    genome editing;    gene knockout;    TALEN;    HMA;   

DOI  :  10.1590/S1415-47572014000100018

来源: SciELO

【 摘 要 】

Engineered nucleases such as zinc finger nucleases (ZFN) and transcription activator-like effector nucleases (TALEN) are one of the most promising tools for modifying genomes. These site-specific enzymes cause double- strand breaks that allow gene disruption or gene insertion, thereby facilitating genetic manipulation. The major problem associated with this approach is the labor-intensive procedures required to screen and confirm the cellular modification by nucleases. In this work, we produced a TALEN that targets the human CCR5 gene and developed a heteroduplex mobility assay for HEK 293T cells to select positive colonies for sequencing. This approach provides a useful tool for the quick detection and easy assessment of nuclease activity.

【 授权许可】

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