Double-stranded RNA-mediated gene suppression
OwnerGeoffrey P. Symonds, Gregory C. Fanning +3inventors
InventorsGeoffrey P. Symonds, Gregory C. Fanning, Gregory M. Arndt, Mitch Raponi, Nham Trieu Tran
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Abstract
The present invention relates to methods for modifying gene expression and in particular to methods for controlling gene expression in eukaryotic cells using double-stranded RNA (dsRNA), and to eukaryotic cell lines in which gene expression has been altered using the method. The invention also relates to compositions suitable for controlling gene expression and to methods of treatment which utilise such compositions.
Claims67
A double-stranded RNA complex comprising:
- (c) a first ribonucleic acid molecule capable of hybridizing under physiological conditions to at least a portion of an mRNA molecule;
- and
- (d) a second ribonucleic acid molecule wherein at least a portion of the second ribonucleic acid molecule is capable of hybridizing under physiological conditions to the first portion.
A linear RNA molecule capable of forming a dsRNA complex wherein the RNA molecule comprises:
- (c) a first portion that hybridizes to at least a portion of a mRNA molecule;
- and
- (d) a second portion wherein at least part of the second portion is capable of hybridizing to the first portion to form a hairpin dsRNA complex.
A linear RNA molecule capable of forming a dsRNA complex wherein the RNA molecule comprises:
- (d) a first portion that comprises a region of RNA that is complementary to at least a portion of a mRNA molecule encoded by a gene;
- (e) a second portion capable of hybridizing to at least part of the first portion;
- and
- (f) a third portion positioned between the first and second portions to permit the first and second portions to hybridize with one another.
A linear RNA molecule capable of forming a dsRNA complex wherein the RNA molecule comprises:
- (c) a first portion that hybridizes to at least a portion of a mRNA molecule encoded by a gene;
- and
- (d) a second portion wherein at least part of the second portion is capable of hybridizing to the first portion and wherein the second portion comprises a polyadenylation signal and a ribozyme positioned between the part of the second portion capable of hybridizing to the first portion and the polyadenylation signal
- wherein the ribozyme is capable of removing the polyadenylation signal.
A method of inhibiting protein expression from a gene in a cell comprising the step of;
- introducing a linear RNA molecule capable of forming a dsRNA complex into a cell, wherein the RNA molecule comprises:
- (a) a first portion that hybridizes to at least part of a mRNA molecule encoded by a gene;
- and
- (b) a second portion wherein at least part of the second portion is capable of hybridizing to the first portion
A method for localizing a dsRNA molecule to the nucleus of a cell comprising the step of:
- introducing a DNA molecule encoding a linear RNA molecule capable of forming a dsRNA complex into a cell wherein the RNA molecule encoded by the DNA molecule comprises:
- (a) a first portion that hybridizes to at least a portion of a mRNA molecule encoded by a gene;
- and
- (b) a second portion wherein at least part of the second portion is capable of hybridizing to the first portion and wherein the second portion comprises a polyadenylation signal and a ribozyme positioned between the part of the second portion capable of hybridizing to the first portion and the polyadenylation signal,
- wherein the ribozyme is capable of removing the polyadenylation signal thereby retaining the RNA in the nucleus.
A method for modulating expression of a nucleic acid sequence in a cell comprising exposing the cell to culture medium in which cells comprising a dsRNA complex comprising a first portion that hybridizes to at least part of a mRNA molecule encoded by a gene;
- and a second portion wherein at least part of the second portion is capable of hybridizing to the first portion have been maintained in cell culture.
A method of identifying the function of a gene in a cell comprising the step of;
- (a) binding a dsRNA molecule to an mRNA molecule in a cell wherein the dsRNA molecule comprises a first ribonucleic acid molecule capable of hybridizing under physiological conditions to at least a portion of an mRNA molecule;
- and a second ribonucleic acid molecule wherein at least a portion of the second ribonucleic acid molecule is capable of hybridizing under physiological conditions to the first portion;
- and
- (b) detecting a change in the cell resulting from the binding.
The composition of claim 61 further comprising a third portion of ribonucleic acid interposed between the first and second portions wherein the third portion promotes hybridization between the first and second portion.
Dependencies are inferred from the claim text.