Rapid selection of tetracycline-controlled inducible cell lines using a green fluorescent-transactivator fusion protein |
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Authors: | Callus B A Mathey-Prevot B |
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Affiliation: | Department of Pediatric Oncology, Dana-Farber Cancer Institute, 44 Binney Street, Boston, Massachusetts, 02115, USA. Callus@dfci.harvard.edu |
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Abstract: | We describe a modification of the tetracycline-controlled expression system that facilitates the rapid identification of tetracycline-sensitive clones. The TetR/VP16 transactivator protein was tagged with the green fluorescent protein (GFP) at its N-terminus. This results in a functional transactivator which allows cells expressing high levels of the modified transactivator to be selected by fluorescent-activated cell sorting. After expansion, single cell clones that maintain a high level of GFP fluorescence can be tested for their ability to transactivate a luciferase gene under control of the Tet operator, leading to the rapid identification of clones with strong inducibility. |
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