Methods of identifying inhibitors of gene silencing in mammalian cells
AssigneeOregon Health & Science University
InventorMitchell Turker
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Abstract
Disclosed herein are methods of identifying inhibitors of gene silencing or re-silencing, which can include repressing expression of a selectable marker gene in mammalian cells, treating the cells with at least one test compound, growing the cells under selective conditions, and quantifying the relative number of cells that live, wherein a change in the relative number of cells as compared to cells that were not treated with the test compound, identifies the compound as an inhibitor of gene silencing or re-silencing. Also disclosed herein are transgenic mice and isolated cell lines that are useful in the disclosed methods and kits for use in performing the disclosed methods.
Claims31
A method of identifying an inhibitor of gene silencing, comprising:
- repressing expression of a selectable marker gene in mammalian cells;
- contacting the cells comprising the repressed selectable marker with at least one test compound;
- growing said cells under selective conditions;
- and
- quantifying cells that live, wherein a change in the relative number of live cells as compared to the relative number of cells that are not treated with the test compound identifies the compound as an inhibitor of gene silencing.
A method for identifying an inhibitor of gene re-silencing, comprising:
- repressing expression of a selectable marker gene in mammalian cells, thereby inducing gene silencing of the selectable marker;
- selecting cells which have undergone reactivation of the selectable marker;
- contacting the cells comprising the reactivated selectable marker with at least one test compound;
- growing said cells under selective conditions;
- and
- quantifying cells that live, wherein a change in the relative number of live cells as compared to the relative number of cells that are not treated with the test compound identifies the compound as an inhibitor of gene re-silencing.
10-12. (canceled)
15-19. (canceled)
23-24. (canceled)
A transgenic mouse comprising a transgenic cell, wherein the cell comprises a first nucleic acid comprising a repressible promoter operably linked to adenine phosphoribosyltransferase (APRT).
30-31. (canceled)
An isolated mouse P19 embryonal carcinoma cell line stably transfected with a tetracycline responsive transcription activator protein operably linked to a constitutive promoter and a selectable marker operably linked to a tetracycline repressible promoter.
(canceled)
A kit for identifying an inhibitor of gene silencing, comprising a cell line stably transfected with a repressible promoter operably linked to a selectable marker gene.
38-41. (canceled)
Dependencies are inferred from the claim text.