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US4870009A
Expired / Lapsed
US
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Method of obtaining gene product through the generation of transgenic animals
Assignee: Salk Institute for Biological Studies
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Filing: 1983-12-15
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Granted: 1989-09-26
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Expires: 2003-12-15
Abstract
Abstract Mammalian genes that encode hormones are cloned and linked to strong promoter DNA sequences. The linked sequences are inserted in plasmids for amplification in prokaryotic cells, and multiple copies of the linked sequences are excised therefrom. Linked sequences are subsequently microinjected into fertilized eggs and the fertilized eggs are implanted into pseudo-pregnent females of the same species. As a result, transgenic animals are born having the linked sequences incorporated into their genomes and expressing the gene-encoded hormone. Because multiple copies of the linked sequences are frequently inserted and because production of the hormone is not limited to certain organs, as is the case with most endogenous hormones, the transgenic animals produce substantial amounts of the hormone. Hormone can be harvested from the living animal (and from its hormone-producing progeny) by extracting fluid, such as blood serum or ascites fluid, on a regular basis.
Inventors
- 👤 Ronald M. Evans
- 👤 Richard D. Palmiter
- 👤 Ralph L. Brinster
Bibliographic Details
Application Number
—
Jurisdiction
US
Data Source
google_patents
Last Synchronized
2026-10-25 15:25
CPC Classifications
C; C07; C07K; C07K14/00; C07K14/435; C07K14/575; C07K14/61; C12; C12N; C12N15/00; C12N15/09; C12N15/63; C12N15/79; C12N15/85; C12N15/87; C12N15/89
Claims Analysis
Total Claims: 17
Claim 1
Independent Claim
145 words
A method of producing a mammalian hormone, which method comprises providing a linerized DNA chain including a promoter/regulator sequences from a mammalian metallothionein gene and a linked sequence that encodes said mammalian hormone under the promotional control of said promoter/regulator sequence, obtaining fertilized mammalian eggs, introducing said linearized DNA chain by microinjection into the male pronuclei of said fertilized mammalian eggs to genetically alter the same, implanting said altered fertilized eggs into host female animals of the species from which said fertilized mammalian eggs were obtained so that the host female animals give birth to animals that develop from said implanted, altered eggs, selecting those of said developed animals which produce said mammalian hormone in amounts very substantially greater than normal, harvesting peripheral blood sera from said selected animals, and isolating said very substantially greater than normal amounts of said hormone from said harvested sera.
Claim 2
Independent Claim
145 words
A method of producing a mammalian hormone, which method comprises providing a linerized DNA chain including a promoter/regulator sequences from a mammalian metallothionein gene and a linked sequence that encodes said mammalian hormone under the promotional control of said promoter/regulator sequence, obtaining fertilized mammalian eggs, introducing said linearized DNA chain by microinjection into the male pronuclei of said fertilized mammalian eggs to genetically alter the same, implanting said altered fertilized eggs into host female animals of the species from which said fertilized mammalian eggs were obtained so that the host female animals give birth to animals that develop from said implanted, altered eggs, selecting those of said developed animals which produce said mammalian hormone in amounts very substantially greater than normal, harvesting peripheral blood sera from said selected animals, and isolating said very substantially greater than normal amounts of said hormone from said harvested sera.
Claim 3
Dependent Claim
30 words
A method according to claim 1 wherein said DNA chain is provided by isolating said promoter/regulator sequence, independently isolating said sequence that encodes a mammalian hormone and linking said sequences.
Claim 4
Dependent Claim
13 words
A method according to claim 1 wherein said second sequence encodes growth hormone.
Claim 5
Dependent Claim
15 words
A method in accordance with claim 3 wherein said second sequence encodes rat growth hormone.
Claim 6
Dependent Claim
14 words
A method according to claim 3 wherein said second sequence encodes human growth hormone.
Claim 7
Dependent Claim
60 words
A method according to claim 1 wherein a said transgenic animals that develop from said fertilized eggs produce an endogenous hormone identical to or analogous to said linked sequence-encoded hormone, said transgenic animal being selected producing said linked sequence-encoded hormone at levels of between about 10 and about 1000 times the level at which it produces the corresponding endogenous hormone.
Claim 8
Dependent Claim
17 words
A method according to claim 1 selecting animals having chromosomes incorporating multiple copies of said DNA chain.
Claim 9
Dependent Claim
21 words
A method according to claim 7 selecting animals that have chromosomes incorporating at least about 5 copies of said DNA chain.
Claim 10
Independent Claim
149 words
A method of producing a mammalian hormone, which method comprises proving a linearized DNA chain including a first sequence which is the promoter/regulator sequence from a mammalian metallothionein gene and a second sequence operably linked to said first sequence that encodes a mammalian hormone, obtaining a fertilized mammalian egg, introducing said linearized DNA chain by microinjection into the male pronuclei of said fertilized mammalian egg to genetically alter the same, implanting said altered fertilized egg into a host female animal of the species from which said fertilized mammalian egg was obtained so that said host female animal gives birth to a transgenic animal that develops from said implanted egg and which produces said hormone in amounts at least 10 times greater than the level at which it produces the corresponding endogenous hormone, harvesting peripheral blood sera from said transgenic animal, and isolating said mammalian hormone from said harvested sera.
Claim 11
Independent Claim
149 words
A method of producing a mammalian hormone, which method comprises proving a linearized DNA chain including a first sequence which is the promoter/regulator sequence from a mammalian metallothionein gene and a second sequence operably linked to said first sequence that encodes a mammalian hormone, obtaining a fertilized mammalian egg, introducing said linearized DNA chain by microinjection into the male pronuclei of said fertilized mammalian egg to genetically alter the same, implanting said altered fertilized egg into a host female animal of the species from which said fertilized mammalian egg was obtained so that said host female animal gives birth to a transgenic animal that develops from said implanted egg and which produces said hormone in amounts at least 10 times greater than the level at which it produces the corresponding endogenous hormone, harvesting peripheral blood sera from said transgenic animal, and isolating said mammalian hormone from said harvested sera.
Claim 12
Dependent Claim
15 words
A method according to claim 9 wherein said promoter/regulator sequence is form a mouse gene.
Claim 13
Dependent Claim
17 words
A method in accordance with claim 9 wherein said promoter/regulator sequence is from a mouse MT-1 gene.
Claim 14
Dependent Claim
14 words
A method according to claim 11 wherein said hormone-encoding sequence encodes rat growth hormone.
Claim 15
Dependent Claim
14 words
A method according to claim 11 wherein said hormone-encoding sequence encodes human growth hormone.
Claim 16
Dependent Claim
15 words
A method according to claim 1 wherein said promoter/regulator sequence is from a mouse gene.
Claim 17
Dependent Claim
18 words
A method in accordance with claim 13 wherein said promoter/ regulator sequence is from a mouse MT-1 gene.
References Cited (Backward Citations)
USPTO Annuity & Maintenance Windows
| Annuity Stage | Window Opens | Payment Deadline | Surcharge Period Until | Estimated Fee | Status |
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| 3.5 Years (Year 4) | 1992-09-25 | 1993-03-27 | 1993-09-26 | $2000.00 | Window Closed / Lapsed |
| 7.5 Years (Year 8) | 1996-09-25 | 1997-03-27 | 1997-09-26 | $3760.00 | Window Closed / Lapsed |
| 11.5 Years (Year 12) | 2000-09-25 | 2001-03-27 | 2001-09-26 | $7700.00 | Window Closed / Lapsed |