Genetic inhibition of double-stranded RNA
View Patent ↗A process is provided of introducing an RNA into a living cell to inhibit gene expression of a target gene in that cell. The process may be practiced ex vivo or in vivo. The RNA has a region with double-stranded structure. Inhibition is sequence-specific in that the nucleotide sequences of the duplex region of the RNA and of a portion of the target gene are identical. The present invention is distinguished from prior art interference in gene expression by antisense or triple-strand methods.
1. A method to inhibit expression of a target gene in an animal cell comprising synthesizing each of a first ribonucleic acid (RNA) and second RNA from an expression construct in the animal cell, wherein the first and second RNAs hybridize to each other to form a double-stranded structure in the cell, the first RNA consisting essentially of a ribonucleotide sequence which corresponds to a nucleotide sequence of the target gene and the second RNA consisting essentially of a ribonucleotide sequence which is complementary to the nucleotide sequence of the target gene, wherein said first and said second RNA are synthesized in the animal cell in an amount sufficient to inhibit the expression of the target gene.
2. A method to inhibit expression of a target gene in an animal cell comprising introducing into said cell double-stranded ribonucleotide (RNA) comprising a first RNA consisting essentially of a ribonucleotide sequence which corresponds to a nucleotide sequence of the target gene and a second RNA which consists essentially of a ribonucleotide sequence which is complementary to the nucleotide sequence of the target gene and wherein said first and second RNAs are present in an amount sufficient to inhibit the expression of the target gene.
3. The method of claim 1 or claim 2 wherein the target gene is an endogenous gene or a transgene.
4. The method of claim 3 wherein the expression of the target gene is inhibited in the cell in vitro.
5. The method of claim 1 or claim 2 wherein the target gene expression is reduced by at least 10%.
6. A method of inhibiting expression of a target gene in an animal cell in a mammal comprising providing at least one ribonucleic acid (RNA) to the animal cell in an amount sufficient to inhibit the expression of a target gene, wherein said RNA is provided to the animal cell by synthesizing said RNA in said animal cell, wherein the RNA comprises or forms a double-stranded structure containing a first strand consisting essentially of a ribonucleotide sequence which corresponds to a nucleotide sequence of the target gene and a second ribonucleotide sequence which is complementary to the target gene, wherein the first and the second ribonucleotide sequences are complementary sequences that hybridize to each other to comprise or form said double-stranded structure, and wherein the RNA comprising or forming the double-stranded structure inhibits expression of target gene.
7. The method of claim 6 , wherein said RNA is transcribed from at least one expression construct.
8. The method of claim 6 , wherein said double-stranded structure is formed by a single self-complementary RNA strand comprising the first and second ribonucleotide sequences.
9. The method of claim 8 , wherein said single self-complementary RNA strand is transcribed from an expression construct.
10. A method of inhibiting the expression of a target gene in a mammalian cell, comprising contacting said mammalian cell with an expression construct, wherein said expression construct comprises an inverted duplication for a segment of the target gene, wherein said segment of the target gene comprises a nucleotide sequence substantially identical to at least one portion of the target gene, wherein a promoter drives expression of said inverted-duplication, and wherein said inverted-duplication forms a double-stranded RNA structure which inhibits expression of the target gene.
11. The method of claim 10 , wherein the double-stranded RNA structure is partially double-stranded.
12. The method of claim 3 , wherein the wherein the expression of the target gene is inhibited in the cell in vivo.
13. The method of claim 1 wherein each of said first and second RNA is synthesized from separate expression constructs.
14. The method of claim 7 wherein said RNA is transcribed from two expression constructs.