Production of fibrinogen in transgenic animals
First Claim
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1. A method for producing biocompetent fibrinogen comprising:
- providing a first DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen Aα
chain, a second DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen Bβ
chain, and a third DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen γ
chain, wherein each chain is from the same species, and wherein each of said first, second and third segments is operably linked to additional DNA segments required for its expression in the mammary gland of a host female mammal;
introducing said DNA segments into a fertilized egg of a non-human mammalian species heterologous to the species of origin of said fibrinogen chains;
inserting said egg into an oviduct or uterus of a female of said mammalian species to obtain offspring carrying said DNA segments;
breeding said offspring to produce female progeny that express said first, second and third DNA segments and produce milk containing biocompetent fibrinogen encoded by said segments;
collecting milk from said female progeny; and
and recovering the biocompetent fibrinogen from the milk.
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Abstract
Materials and methods for producing fibrinogen in transgenic non-human mammals are disclosed. DNA segments encoding Aα, Bβ and γ chains of fibrinogen are introduced into the germ line of a non-human mammal, and the mammal or its female progeny produces milk containing fibrinogen expressed from the introduced DNA segments. Non-human mammalian embryos and transgenic non-human mammals carrying DNA segments encoding heterologous fibrinogen polypeptide chains are also disclosed.
45 Citations
33 Claims
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1. A method for producing biocompetent fibrinogen comprising:
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providing a first DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen Aα
chain, a second DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen Bβ
chain, and a third DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen γ
chain, wherein each chain is from the same species, and wherein each of said first, second and third segments is operably linked to additional DNA segments required for its expression in the mammary gland of a host female mammal;introducing said DNA segments into a fertilized egg of a non-human mammalian species heterologous to the species of origin of said fibrinogen chains; inserting said egg into an oviduct or uterus of a female of said mammalian species to obtain offspring carrying said DNA segments; breeding said offspring to produce female progeny that express said first, second and third DNA segments and produce milk containing biocompetent fibrinogen encoded by said segments; collecting milk from said female progeny; and and recovering the biocompetent fibrinogen from the milk. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11)
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12. A method of producing biocompetent fibrinogen comprising:
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incorporating a first DNA segment encoding a secretion signal operably linked to an Aα
chain of fibrinogen into a β
-lactoglobulin gene to produce a first gene fusion comprising a β
-lactoglobulin promoter operably linked to the first DNA segment;incorporating a second DNA segment encoding a secretion signal operably linked to a Bβ
chain of fibrinogen into a β
-lactoglobulin gene to produce a second gene fusion comprising a β
-lactoglobulin promoter operably linked to the second DNA segment;incorporating a third DNA segment encoding a secretion signal operably linked to a γ
chain of fibrinogen into a β
-lactoglobulin gene to produce a third gene fusion comprising a β
-lactoglobulin promoter operably linked to the third DNA segment wherein each of said first, second and third segments are of the same species;introducing said first, second and third gene fusions into the germ line of a non-human mammal so that said DNA segments are expressed in a mammary gland of said mammal or its female progeny and biocompetent fibrinogen is secreted into milk of said mammal or its female progeny; obtaining milk from said mammal or its female progeny; and recovering said fibrinogen from said milk. - View Dependent Claims (13, 14, 15, 16, 17)
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18. A method for producing biocompetent fibrinogen comprising:
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providing a transgenic female non-human mammal carrying in its germline heterologous DNA segments encoding Aα
, Bβ and
γ
chains of fibrinogen, wherein said segments are expressed in a mammary gland of said mammal and biocompetent fibrinogen encoded by said segments is secreted into milk of said mammal;collecting milk from said mammal; and recovering said biocompetent fibrinogen from said milk. - View Dependent Claims (19, 20)
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21. A transgenic non-human female mammal that produces recoverable amounts of biocompetent human fibrinogen in its milk, wherein said mammal comprises:
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a first DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen Aα
chain,a second DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen Bβ
chain, anda third DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen γ
chain, andfurther wherein each chain is derived from the same species and is operably linked to additional DNA segments required for its expression in the mammary gland of a host female mammal. - View Dependent Claims (22)
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23. A process for producing a transgenic offspring of a mammal comprising:
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providing a first DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen Aα
chain, a second DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen Bβ
chain, and a third DNA segment encoding a secretion signal operably linked to a heterologous fibrinogen γ
chain, wherein each chain is derived from the same species, and wherein each of said first, second and third segments is operably linked to additional DNA segments required for its expression in the mammary gland of a host female mammal;introducing said DNA segments into a fertilized egg of a non-human mammalian species heterologous to the species of origin of said fibrinogen chains; inserting said fertilized egg into an oviduct or uterus of a female of said mammalian species; and allowing said fertilized egg to develop thereby producing transgenic offspring carrying said first, second and third DNA segments, wherein female progeny of said mammal express said DNA segments in a mammary gland to produce biocompetent fibrinogen. - View Dependent Claims (24, 25, 26, 27, 28, 29)
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30. A non-human mammal carrying in its germline DNA segments encoding human Aα
- , Bβ and
γ
chains of fibrinogen, wherein female progeny of said mammal express said DNA segments in a mammary gland to produce biocompetent human fibrinogen. - View Dependent Claims (31, 32, 33)
- , Bβ and
Specification