US5550035A
Expired / Lapsed
US
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Prokaryotic expression in eukaryotic cells
Assignee: Research Foundation of the State University of New York
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Filing: 1994-01-26
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Granted: 1996-08-27
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Expires: 2014-01-26
Abstract
Abstract A transient expression system is disclosed that utilizes bacteriophage RNA polymerase in the presence of a DNA-based cytoplasmic virus to facilitate expression of a foreign gene in the cytoplasm of a eukaryotic cell. A method of expressing a foreign gene in the cytoplasm of a eukaryotic cell is also disclosed which comprises incorporating into the cytoplasm a DNA-based cytoplasmic virus, a suitable carrier comprising a gene for an RNA polymerase which gene is foreign to the carrier and to the cells, and a suitable carrier comprising a functional, cistron including a foreign gene flanked by a promotor sequence which is recognized by the RNA polymerase.
Inventors
- 👤 Bernard Moss
- 👤 F. William Studier
- 👤 Thomas R. Fuerst
- 👤 Edward G. Niles
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-11-01 20:31
CPC Classifications
C; C12; C12N; C12N9/00; C12N9/10; C12N9/12; C12N9/1241; C12N9/1247; C12N15/00; C12N15/09; C12N15/10; C12N15/63; C12N15/79; C12N15/85
Claims Analysis
Total Claims: 47
Claim 1
Independent Claim
114 words
A method of expressing a gene in a eukaryotic cell comprising incorporating into said cell: a DNA-based cytoplasmic virus; a first carrier including a first gene encoding a bacteriophage RNA polymerase selected from the group consisting of T7, SP6, GH1 and T3 RNA polymerase, said first gene being foreign to the carrier and to the cell, and a first promoter sequence which initiates expression of said first gene; and a second carrier comprising a functional cistron comprising a second foreign gene including a second promoter sequence responsive to said bacteriophage RNA polymerase, such that upon recognition of said RNA polymerase, said second promoter initiates expression of the second foreign gene in the eukaryotic cell.
Claim 2
Independent Claim
114 words
A method of expressing a gene in a eukaryotic cell comprising incorporating into said cell: a DNA-based cytoplasmic virus; a first carrier including a first gene encoding a bacteriophage RNA polymerase selected from the group consisting of T7, SP6, GH1 and T3 RNA polymerase, said first gene being foreign to the carrier and to the cell, and a first promoter sequence which initiates expression of said first gene; and a second carrier comprising a functional cistron comprising a second foreign gene including a second promoter sequence responsive to said bacteriophage RNA polymerase, such that upon recognition of said RNA polymerase, said second promoter initiates expression of the second foreign gene in the eukaryotic cell.
Claim 3
Dependent Claim
18 words
The method of claim 1, wherein said virus is selected from the group consisting of poxvirus and iridovirus.
Claim 4
Dependent Claim
10 words
The method of claim 2, wherein said virus is poxvirus.
Claim 5
Dependent Claim
24 words
The method of claim 3, wherein said poxvirus is selected from the group consisting of orthopoxvirus, parapoxvirus, avipoxvirus, capripoxvirus, leporipoxvirus, suipoxvirus, molluscipoxvirus and yatapoxvirus.
Claim 6
Dependent Claim
10 words
The method of claim 4, wherein said poxvirus is orthopoxvirus.
Claim 7
Dependent Claim
10 words
The method of claim 5, wherein said orthopoxvirus is vaccinia.
Claim 8
Dependent Claim
12 words
The method of claim 1, wherein said first carrier is a plasmid.
Claim 9
Dependent Claim
13 words
The method of claim 7, wherein said first promoter is a poxvirus promoter.
Claim 10
Dependent Claim
15 words
The method of claim 8, wherein said first gene is the T7 RNA polymerase gene.
Claim 11
Dependent Claim
15 words
The method of claim 9, wherein said plasmid is pTF7-3 having A.T.C.C. Designation Number 67202.
Claim 12
Dependent Claim
12 words
The method of claim 1, wherein said second carrier is a plasmid.
Claim 13
Dependent Claim
13 words
The method of claim 1, wherein said second promoter is a T7 promoter.
Claim 14
Dependent Claim
14 words
The method of claim 11, wherein said second foreign gene is a eukaryotic gene.
Claim 15
Dependent Claim
13 words
The method of claim 11, wherein said cistron further includes a terminator sequence.
Claim 16
Independent Claim
110 words
A method of expressing a gene in a eukaryotic cell comprising incorporating into said cell: a DNA-based cytoplasmic virus including a first gene encoding a bacteriophage RNA polymerase selected from the group consisting of T7, SP6, GH1 and T3 RNA polymerase, which first gene is foreign to the virus and to the cell, and a first promoter sequence which initiates expression of the first gene; and a carrier comprising a functional cistron comprising a second foreign gene including a second promoter sequence responsive to said bacteriophage RNA polymerase, such that upon recognition of said RNA polymerase, said second promoter initiates expression of the second foreign gene in the eukaryotic cell.
Claim 17
Independent Claim
110 words
A method of expressing a gene in a eukaryotic cell comprising incorporating into said cell: a DNA-based cytoplasmic virus including a first gene encoding a bacteriophage RNA polymerase selected from the group consisting of T7, SP6, GH1 and T3 RNA polymerase, which first gene is foreign to the virus and to the cell, and a first promoter sequence which initiates expression of the first gene; and a carrier comprising a functional cistron comprising a second foreign gene including a second promoter sequence responsive to said bacteriophage RNA polymerase, such that upon recognition of said RNA polymerase, said second promoter initiates expression of the second foreign gene in the eukaryotic cell.
Claim 18
Dependent Claim
10 words
The method of claim 15, wherein said virus is poxvirus.
Claim 19
Dependent Claim
10 words
The method of claim 16, wherein said poxvirus is vaccinia.
Claim 20
Dependent Claim
16 words
The method of claim 15, wherein said virus is vTF7-3 having A.T.C.C. Designation Number VR 2153.
Claim 21
Dependent Claim
13 words
The method of claim 15, wherein said carrier is a DNA-based cytoplasmic virus.
Claim 22
Dependent Claim
15 words
The method of claim 19, wherein said first gene is the T7 RNA polymerase gene.
Claim 23
Dependent Claim
13 words
The method of claim 15, wherein said second promoter is a T7 promoter.
Claim 24
Dependent Claim
11 words
The method of claim 15, wherein said carrier is a plasmid.
Claim 25
Dependent Claim
10 words
The method of claim 22, wherein said plasmid is pTF7CAT-1.
Claim 26
Dependent Claim
10 words
The method of claim 22, wherein said virus is poxvirus.
Claim 27
Dependent Claim
10 words
The method of claim 24, wherein said poxvirus is vaccinia.
Claim 28
Dependent Claim
10 words
The method of claim 22, wherein said plasmid is PTF7LZ-1.
Claim 29
Dependent Claim
16 words
The method of claim 19, wherein said carrier is vTF7LZ-1 having A.T.C.C. Designation Number VR 2152.
Claim 30
Dependent Claim
16 words
The method of claim 19, wherein said carrier is vTF7HB-1 having A.T.C.C. Designation Number VR 2154.
Claim 31
Dependent Claim
12 words
The method of claim 22, wherein said second foreign gene encodes HBsAg.
Claim 32
Dependent Claim
13 words
The method of claim 29, wherein said second promoter is a T7 promoter.
Claim 33
Dependent Claim
12 words
The method of claim 29, wherein said second foreign gene is lacZ.
Claim 34
Dependent Claim
12 words
The method of claim 29, wherein said second foreign gene is CAT.
Claim 35
Dependent Claim
115 words
A eukaryotic cell capable of expressing a foreign gene in the cytoplasm of the cell, comprising incorporated into said cell: a DNA-based cytoplasmic virus including a first gene encoding a bacteriophage RNA polymerase selected from the group consisting of T7, SP6, GH1 and T3 RNA polymerase, said first gene being foreign to the virus and to the cell, and a first promoter sequence which initiates expression of said first gene; and a carrier comprising a functional cistron comprising a second foreign gene including a second promoter sequence responsive to said bacteriophage RNA polymerase, such that upon recognition of said RNA polymerase, said second promoter initiates expression of the second foreign gene in the eukaryotic cell.
Claim 36
Dependent Claim
115 words
A eukaryotic cell capable of expressing a foreign gene in the cytoplasm of the cell, comprising incorporated into said cell: a DNA-based cytoplasmic virus including a first gene encoding a bacteriophage RNA polymerase selected from the group consisting of T7, SP6, GH1 and T3 RNA polymerase, said first gene being foreign to the virus and to the cell, and a first promoter sequence which initiates expression of said first gene; and a carrier comprising a functional cistron comprising a second foreign gene including a second promoter sequence responsive to said bacteriophage RNA polymerase, such that upon recognition of said RNA polymerase, said second promoter initiates expression of the second foreign gene in the eukaryotic cell.
Claim 37
Dependent Claim
10 words
The cell of claim 33, wherein said virus is vaccinia.
Claim 38
Dependent Claim
15 words
The cell of claim 33, wherein said first gene is the T7 RNA polymerase gene.
Claim 39
Dependent Claim
11 words
The cell of claim 33, wherein said carrier is a plasmid.
Claim 40
Dependent Claim
13 words
The cell of claim 33, wherein said second promoter is a T7 promoter.
Claim 41
Dependent Claim
13 words
The cell of claim 33, wherein said cistron further includes a terminator sequence.
Claim 42
Dependent Claim
13 words
The cell of claim 33, wherein said carrier is a DNA-based cytoplasmic virus.
Claim 43
Dependent Claim
10 words
The cell of claim 39, wherein said carrier is vaccina.
Claim 44
Dependent Claim
13 words
The cell of claim 39, wherein said second promoter is a T7 promoter.
Claim 45
Dependent Claim
13 words
The cell of claim 39, wherein said cistron further includes a terminator sequence.
Claim 46
Dependent Claim
26 words
A DNA-based cytoplasmic virus containing a foreign gene encoding a bacteriophage RNA polymerase selected from the group consisting of T7, SP6, GH1 and T3 RNA polymerase.
Claim 47
Dependent Claim
26 words
A DNA-based cytoplasmic virus containing a foreign gene encoding a bacteriophage RNA polymerase selected from the group consisting of T7, SP6, GH1 and T3 RNA polymerase.
References Cited (Backward Citations)
US5126251A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1999-08-27 | 2000-02-26 | 2000-08-27 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 2003-08-27 | 2004-02-26 | 2004-08-27 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2007-08-27 | 2008-02-26 | 2008-08-27 | $7700.00 | Window Closed / Lapsed |