US8183217B2
Expired / Lapsed
US
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Methods and means for obtaining modified phenotypes
Assignee: Commonwealth Scientific and Industrial Research Organisation
•
Filing: 2005-07-13
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Granted: 2012-05-22
•
Expires: 2025-07-13
Abstract
Abstract Methods and means are provided for reducing the phenotypic expression of a nucleic acid of interest in eukaryotic cells by providing aberrant, preferably unpolyadenylated, target-specific RNA to the nucleus of the host cell. Preferably, the unpolyadenylated, target-specific RNA is provided by transcription of a chimeric gene comprising a promoter and a DNA region encoding the target-specific RNA.
Inventors
- 👤 Peter Michael Waterhouse
- 👤 Ming-Bo Wang
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-07 13:41
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/63; C12N15/79; C12N15/82; C12N15/8216; C12N15/8218; C12N15/8241; C12N15/8242; C12N15/8243; C12N15/8247; C12N15/8261; C12N15/8271; C12N15/8279; C12N15/8283; C12N9/00; C12N9/14; C12N9/24; C12N9/2402; C12N9/2405; C12Y; C12Y302/00; C12Y302/01; C12Y302/01031
Claims Analysis
Total Claims: 21
Claim 1
Independent Claim
176 words
A method for expressing an unpolyadenylated hairpin RNA in an animal cell, the method comprising the step of providing to an animal cell in culture a DNA comprising a promoter operably linked to a target specific DNA region encoding the unpolyadenylated hairpin RNA, wherein the unpolyadenylated hairpin RNA comprises a target specific sense nucleotide sequence and a target specific antisense nucleotide sequence, wherein the target specific antisense nucleotide sequence consists of about 20 consecutive nucleotides in a sequence identical to the sequence of the complement of a part of an RNA molecule transcribed or produced from a nucleic acid of interest in the animal cell, wherein the target specific sense nucleotide sequence consists of about 20 consecutive nucleotides in a sequence identical to the sequence of the part of the RNA molecule transcribed or produced from the nucleic acid of interest, and wherein the target specific sense nucleotide sequence and the target specific antisense nucleotide sequence are complementary to each other, so as to form in the nucleus of the animal cell the unpolyadenylated hairpin RNA.
Claim 2
Independent Claim
176 words
A method for expressing an unpolyadenylated hairpin RNA in an animal cell, the method comprising the step of providing to an animal cell in culture a DNA comprising a promoter operably linked to a target specific DNA region encoding the unpolyadenylated hairpin RNA, wherein the unpolyadenylated hairpin RNA comprises a target specific sense nucleotide sequence and a target specific antisense nucleotide sequence, wherein the target specific antisense nucleotide sequence consists of about 20 consecutive nucleotides in a sequence identical to the sequence of the complement of a part of an RNA molecule transcribed or produced from a nucleic acid of interest in the animal cell, wherein the target specific sense nucleotide sequence consists of about 20 consecutive nucleotides in a sequence identical to the sequence of the part of the RNA molecule transcribed or produced from the nucleic acid of interest, and wherein the target specific sense nucleotide sequence and the target specific antisense nucleotide sequence are complementary to each other, so as to form in the nucleus of the animal cell the unpolyadenylated hairpin RNA.
Claim 3
Dependent Claim
26 words
The method of claim 1 , wherein the target specific sense nucleotide sequence corresponds to one or more consecutive exons of the nucleic acid of interest.
Claim 4
Dependent Claim
24 words
The method of claim 1 , wherein the target specific sense nucleotide sequence corresponds to a translated region of the nucleic acid of interest.
Claim 5
Dependent Claim
28 words
The method of claim 1 , wherein the target specific sense nucleotide sequence corresponds to an untranslated region of the RNA produced from the nucleic acid of interest.
Claim 6
Dependent Claim
32 words
The method of claim 1 , wherein the promoter is recognized by a eukaryotic RNA polymerase I or III and the DNA further comprises a terminator for the polymerase I or III.
Claim 7
Dependent Claim
23 words
The method of claim 1 , wherein the nucleic acid of interest is a gene incorporated in the genome of the animal cell.
Claim 8
Dependent Claim
20 words
The method of claim 1 , wherein the nucleic acid of interest is an endogenous gene of the animal cell.
Claim 9
Dependent Claim
17 words
The method of claim 1 , wherein the nucleic acid of interest is a viral nucleic acid.
Claim 10
Dependent Claim
15 words
The method of claim 1 , wherein the unpolyadenylated RNA lacks a 5′ cap structure.
Claim 11
Dependent Claim
59 words
The method of claim 1 , comprising the steps of a) introducing a DNA into the nucleus of the animal cell, the DNA encoding the unpolyadenylated hairpin RNA, and b) observing a phenotype of the animal cell by a suitable method; thereby identifying a phenotype associated with the expression of the nucleic acid of interest in the animal cell.
Claim 12
Dependent Claim
17 words
The method of claim 10 , wherein the phenotype is observed after culturing of the animal cell.
Claim 13
Dependent Claim
20 words
The method of claim 10 , wherein the phenotype is a modified profile of metabolites synthesized in the animal cell.
Claim 14
Dependent Claim
14 words
The method of claim 1 , wherein the animal cell is a human cell.
Claim 15
Dependent Claim
13 words
The method of claim 1 , wherein the promoter is a constitutive promoter.
Claim 16
Dependent Claim
13 words
The method of claim 1 , wherein the promoter is an inducible promoter.
Claim 17
Dependent Claim
13 words
The method of claim 1 , wherein the promoter is a tissue-specific promoter.
Claim 18
Dependent Claim
20 words
The method of claim 1 , wherein the promoter is recognized by a single subunit RNA polymerase from a bacteriophage.
Claim 19
Dependent Claim
23 words
The method of claim 1 , wherein the nucleic acid of interest is a transgene that has been introduced into the animal cell.
Claim 20
Dependent Claim
15 words
The method of claim 1 , wherein the unpolyadenylated hairpin RNA comprises a persistent intron.
Claim 21
Dependent Claim
28 words
The method of claim 5 , wherein the promoter is recognized by a eukaryotic RNA polymerase III and the DNA further comprises a terminator for the polymerase III.
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