US4766067A
Expired / Lapsed
US
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Gene amplification
Assignee: Harvard University
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Filing: 1985-05-31
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Granted: 1988-08-23
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Expires: 2005-05-31
Abstract
Abstract Hybrid DNA capable of effecting controlled amplification in a host cell of a heterologous gene that determines a specific function. The hybrid DNA has the following three different regions of DNA: (1) an "amplicon" that is inducible by an external stimulus; (2) the heterologous gene (or a site for its insertion); and (3) a selectable marker enabling isolation of the transformed host cells. In order to make a desired compound coded by the heterologous gene, a vehicle including the hybrid DNA is introduced into a host cell is transformed with the vector, transformed host cells are grown to a desired population, the external stimulus is controlled to effect amplification of the gene in members of the population, cells containing amplified gene are cultured, and the compound is recovered.
Inventors
- 👤 Debajit K. Biswas
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-24 16:32
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/63; C12N15/79; C12N15/85; C07; C07K; C07K14/00; C07K14/435; C07K14/575; C07K14/61; C12N15/67
Claims Analysis
Total Claims: 13
Claim 1
Independent Claim
96 words
Hybrid DNA capable of effecting the amplification in a host cell of a heterologous gene determining a desired function, said hybrid DNA comprising two regions: (1) a first DNA region comprising an amplicon inducible by 5-bromodeoxyuridine, said amplicon being a segment from the 10.3 kb rat prolactin amplicon-containing fragment of ATCC deposit 40187, and (2) a second DNA region adjacent to said first DNA region comprising said heterologous gene, said hybrid DNA being capable of amplifying the number of copies of said heterologous gene in a host cell transformed with said hybrid DNA responsive to 5-bromodeoxyuridine.
Claim 2
Dependent Claim
96 words
Hybrid DNA capable of effecting the amplification in a host cell of a heterologous gene determining a desired function, said hybrid DNA comprising two regions: (1) a first DNA region comprising an amplicon inducible by 5-bromodeoxyuridine, said amplicon being a segment from the 10.3 kb rat prolactin amplicon-containing fragment of ATCC deposit 40187, and (2) a second DNA region adjacent to said first DNA region comprising said heterologous gene, said hybrid DNA being capable of amplifying the number of copies of said heterologous gene in a host cell transformed with said hybrid DNA responsive to 5-bromodeoxyuridine.
Claim 3
Dependent Claim
20 words
The hybrid DNA of claim 1 further comprising a selectable marker wherein said selectable marker is a thymidine kinase gene.
Claim 4
Dependent Claim
21 words
The hybrid DNA of claim 1 further comprising a selectable marker wherein said selectable marker gene encodes resistance to an antibiotic.
Claim 5
Dependent Claim
15 words
The hybrid DNA of claim 1 wherein said heterologous gene codes for human growth hormone.
Claim 6
Dependent Claim
9 words
A cell comprising the hybrid DNA of claim 1.
Claim 7
Dependent Claim
12 words
The cell of claim 5 wherein said cell is a eucaryotic cell.
Claim 8
Dependent Claim
13 words
The cell of claim 6 wherein said eucaryotic cell is a mammalian cell.
Claim 9
Independent Claim
150 words
A method of producing a desired compound comprising, introducing into a host cell hybrid DNA capable of effecting the amplification in a host cell of a heterologous gene determining a desired function said hybrid DNA comprising two regions: (1) a first DNA region comprising an amplicon inducible by 5-bromodeoxyuridine, said amplicon being a segment from the 10.3 kb rat prolactin amplicon-containing fragment of ATCC deposit 40187, and (2) a second DNA region adjacent to said first DNA region comprising said heterologous gene, said hybrid DNA being capable of amplifying the number of copies of said heterologous gene in a host cell transformed with said hybrid DNA responsive to 5-bromodeoxyuridine, growing said host cells with said hybrid DNA to a desired population, changing an external stimulus to effect amplification of said heterologous gene in the members of said population, culturing cells in which said gene is amplified, and recovering said compound.
Claim 10
Independent Claim
150 words
A method of producing a desired compound comprising, introducing into a host cell hybrid DNA capable of effecting the amplification in a host cell of a heterologous gene determining a desired function said hybrid DNA comprising two regions: (1) a first DNA region comprising an amplicon inducible by 5-bromodeoxyuridine, said amplicon being a segment from the 10.3 kb rat prolactin amplicon-containing fragment of ATCC deposit 40187, and (2) a second DNA region adjacent to said first DNA region comprising said heterologous gene, said hybrid DNA being capable of amplifying the number of copies of said heterologous gene in a host cell transformed with said hybrid DNA responsive to 5-bromodeoxyuridine, growing said host cells with said hybrid DNA to a desired population, changing an external stimulus to effect amplification of said heterologous gene in the members of said population, culturing cells in which said gene is amplified, and recovering said compound.
Claim 11
Dependent Claim
12 words
The method of claim 8 wherein said host cell is a eukaryote.
Claim 12
Dependent Claim
14 words
The method of claim 9 wherein said host encaryotic cell is a mammalian cell.
Claim 13
Dependent Claim
12 words
The method of claim 8 wherein said compound is human growth hormone.
References Cited (Backward Citations)
US4656134A
Cited
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
|---|---|---|---|---|---|
| 3.5 Years (Year 4) | 1991-08-23 | 1992-02-22 | 1992-08-23 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1995-08-23 | 1996-02-22 | 1996-08-23 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 1999-08-23 | 2000-02-22 | 2000-08-23 | $7700.00 | Window Closed / Lapsed |