Bistable genetic toggle switch
AssigneeCellicon Technologies, Inc.
InventorsTimothy Gardner, James J. Collins
Life of this patent
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Abstract
Provided are methods and compositions for regulating gene expression in a cell. The invention provides recombinant genetic toggle switches which contain a first constitutive promoter-regulatory gene operon and a second constitutive promoter-regulatory gene operon. Expression of the regulatory gene from the first operon inhibits expression from the promoter in the second operon, and expression of the regulatory gene from the second operon inhibits expression from the promoter in the first operon. By use of the toggle switch and various switching agents it is possible to reversibly switch the expression of a gene of interest between a stable “on” state and stable “off” state or vice versa via transient exposure to a switching agent.
Claims30
A recombinant bistable genetic toggle switch that is capable of being stable in a first state or in a second state, the toggle switch comprising:
- (a) a first nucleic acid construct comprising a first constitutive promoter operably associated with a first gene encoding a first repressor protein;
- and
- (b) a second nucleic acid construct comprising a second constitutive promoter operably associated with a second gene encoding a second repressor protein,
- wherein the second repressor protein, when produced, is capable of repressing transcription from the first promoter, and wherein repression of the first promoter by the second repressor protein is reducible by a first switching agent, and
- wherein the first repressor protein, when produced, is capable of repressing transcription from the second promoter, and wherein repression of the second promoter by the first repressor protein is reducible by a second switching agent.
A method of alternating transcription from first and second promoters in a host cell, the method comprising the steps of:
- (i) providing a host cell harboring a recombinant bistable genetic switch comprising:
- (a) a first nucleic acid construct comprising a first constitutive promoter operably associated with a first gene encoding a first repressor protein;
- and
- (b) a second nucleic acid construct comprising a second constitutive promoter operably associated with a second gene encoding a second repressor protein,
- wherein the second repressor protein, when produced, is capable of repressing transcription from the first promoter, and wherein repression of the first promoter by the second repressor protein is reduced by a first switching agent, and
- wherein the first repressor protein, when produced, is capable of repressing transcription from the second promoter, and wherein repression of the second promoter by the first repressor protein is reduced by a second switching agent;
- and
- (ii) providing either the first switching agent to derepress transcription of the first gene by the first promoter or the second switching agent to derepress transcription of the second gene by the second promoter.
Dependencies are inferred from the claim text.