Inactivation of an Integrated Antibiotic Resistance Gene in Mammalian Cells to Re-Enable Antibiotic Selection | Zendy

Peiling Ni | Zendy; Qian Zhang | Zendy; Hạixia Chen | Zendy; Lingyi Chen | Zendy

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Inactivation of an Integrated Antibiotic Resistance Gene in Mammalian Cells to Re-Enable Antibiotic Selection

Author(s) -

Peiling Ni,

Qian Zhang,

Hạixia Chen,

Lingyi Chen

Publication year - 2014

Publication title -

biotechniques

Language(s) - English

Resource type - Journals

SCImago Journal Rank - 0.617

H-Index - 131

eISSN - 1940-9818

pISSN - 0736-6205

DOI - 10.2144/000114160

Subject(s) - puromycin , crispr , gene , biology , embryonic stem cell , antibiotics , antibiotic resistance , reporter gene , gene targeting , cell culture , genetics , microbiology and biotechnology , gene expression , protein biosynthesis

Removing an antibiotic resistance gene allows the same antibiotic to be re-used in the next round of genetic manipulation. Here we applied the CRISPR/Cas system to disrupt the puromycin resistance gene in an engineered mouse embryonic stem cell line and then re-used puromycin selection in the resulting cells to establish stable reporter cell lines. With the CRISPR/Cas system, pre-engineered sequences, such as loxP or FRT, are not required. Thus, this technique can be used to disrupt antibiotic resistance genes that cannot be removed by the Cre-loxP and Flp-FRT systems.

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