US4740461A
Expired / Lapsed
US
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Vectors and methods for transformation of eucaryotic cells
Assignee: Genetics Institute LLC
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Filing: 1983-12-27
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Granted: 1988-04-26
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Expires: 2003-12-27
Abstract
Abstract Eucaryotic cells cotransformed with product and selection genes yield considerably greater quantities of product after a novel subcloning strategy is employed: Transformants are identified for product yield, cultured under selection pressure and the progeny screened for product yield. Novel transformation vectors contain directly ligated selection and product genes and/or eucaryotic promoters.
Inventors
- 👤 Randal J. Kaufman
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-29 13:51
CPC Classifications
C; C12; C12N; C12N9/00; C12N9/14; C12N9/48; C12N9/50; C12N9/64; C12N9/6421; C12N9/6424; C12N9/6456; C12N9/6459; C12N15/00; C12N15/09; C12N15/63; C12N15/79; C12N15/85; C12Y; C12Y304/00; C12Y304/21; C12Y304/21069
Claims Analysis
Total Claims: 13
Claim 1
Independent Claim
101 words
In a method for transforming eukaryotic cells to incorporate a desired product gene by the technique of cotransfecting with unlinked vectors, one of which contains said product gene, the improvement comprising: cotransfecting the cells with (a) a first vector comprising an enhancer, a first promoter, and a product gene under the transciptional control of the first promoter, said first vector not containing a selection gene under the control of the first promoter; and, (b) a second unlinked vector comprising a selection gene under the transciptional control of a second promoter dependant on the enhancer, the second vector not containing the enhancer.
Claim 2
Dependent Claim
101 words
In a method for transforming eukaryotic cells to incorporate a desired product gene by the technique of cotransfecting with unlinked vectors, one of which contains said product gene, the improvement comprising: cotransfecting the cells with (a) a first vector comprising an enhancer, a first promoter, and a product gene under the transciptional control of the first promoter, said first vector not containing a selection gene under the control of the first promoter; and, (b) a second unlinked vector comprising a selection gene under the transciptional control of a second promoter dependant on the enhancer, the second vector not containing the enhancer.
Claim 3
Dependent Claim
34 words
A method of claim 1 which further comprises selecting the cotransfected cells so produced which are capable of expressing both the product and selection genes and separating the cells so selected from other cells.
Claim 4
Dependent Claim
12 words
Transformed cells produced by the method of claim 2 or progeny thereof.
Claim 5
Dependent Claim
46 words
A method for producing a protein product which comprises the steps of culturing a transformed cell of claim 3 in a suitable culture medium and under suitable conditions permitting the expression of the product gene, and recovering the protein product from the medium and the cells.
Claim 6
Dependent Claim
21 words
A method of claim 1, wherein each vector also contains a polyadenylation site downstream of the product and selection genes, respectively.
Claim 7
Dependent Claim
12 words
Transformed cells produced by the method of claim 5 or progeny thereof.
Claim 8
Dependent Claim
46 words
A method for producing a protein product which comprises the steps of culturing a transformed cell of claim 6 in a suitable culture medium and under suitable conditions permitting the expression of the product gene, and recovering the protein product from the medium and the cells.
Claim 9
Dependent Claim
15 words
A method of claim 1, wherein the product gene encodes a human tissue plasminogen activator.
Claim 10
Dependent Claim
12 words
Transformed cells produced by the method of claim 8 or progeny thereof.
Claim 11
Dependent Claim
56 words
A method for producing a human tissue plasminogen activator which comprises the steps of culturing a transformed cell of claim 9 in a suitable culture medium and under suitable conditions permitting the expression of the product gene encoding the human tissue plasminogen activator, and recovering the human tissue plasminogen activator from the medium and the cells.
Claim 12
Dependent Claim
12 words
Transformed cells produced by the method of claim 1 or progeny thereof.
Claim 13
Dependent Claim
46 words
A method for producing a protein product which comprises the steps of culturing a transformed cell of claim 11 in a suitable culture medium and under suitable conditions permitting the expression of the product gene, and recovering the protein product from the medium and the cells.
References Cited (Backward Citations)
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1991-04-26 | 1991-10-26 | 1992-04-26 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1995-04-26 | 1995-10-26 | 1996-04-26 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1999-04-26 | 1999-10-26 | 2000-04-26 | $7700.00 | Window Closed / Lapsed |