Transgenic animals as urinary bioreactors for the production of polypeptide in the urine, recombinant DNA construct for kidney-specific expression, and method of using same
AssigneeNew York University
InventorsXue-Ru Wu, Tung-Tien Sun
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Abstract
The invention relates to recombinant DNA constructs, a method for producing a recombinant biologically active protein in vivo in the urine of a non-human mammal using a kidney-specific promoter, such as the uromodulin promoter, and the transgenic non-human mammals that serve as urine-based bioreactors for protein production.
Claims26
A method for producing a recombinant biologically active polypeptide, comprising:
- introducing an isolated DNA molecule comprising a mammalian uromodulin promoter operably linked to a heterologous DNA sequence encoding a heterologous polypeptide, wherein said mammalian uromodulin promoter directs expression of said heterologous polypeptide in vivo to the ascending limb of Henle's loop and the early distal tubules of the kidneys to produce a recombinant biologically active polypeptide in the urine, into a fertilized embryo of a non-human mammal selected from the group consisting of goat, cattle, sheep, pig, and mouse to generate a transgenic non-human mammal which expresses and secretes the heterologous polypeptide into the urine of the transgenic non-human mammal as a recombinant biologically active polypeptide;
- collecting urine from the transgenic non-human mammal;
- and
- recovering the secreted polypeptide to produce a recombinant biologically active polypeptide.
A transgenic non-human mammal all of whose germ cells and somatic cells contain a recombinant construct corresponding to a DNA molecule comprising a mammalian uromodulin promoter operably linked to a heterologous DNA sequence encoding a heterologous polypeptide, wherein:
- said mammalian uromodulin promoter directs expression of said heterologous polypeptide in vivo to the ascending limb of Henle's loop and the early distal tubules of the kidneys to produce a recombinant biologically active polypeptide in the urine
- said DNA molecule has been introduced into said mammal selected from the group consisting of goat, sheep, cattle, pig and mouse, at an embryonic stage;
- and
- said mammal produces recoverable amounts of a recombinant biologically active polypeptide in its urine.
A method for producing a recombinant biologically active polypeptide, comprising:
- introducing an isolated DNA molecule comprising a mammalian uromodulin promoter, selected from the group consisting of mouse, goat, human, rat, and bovine uromodulin promoters and operably linked to a heterologous DNA sequence encoding a heterologous polypeptide, into a fertilized embryo of a non-human mammal selected from the group consisting of goat, cattle, sheep, pig and mouse to generate a transgenic non-human mammal which expresses said heterologous polypeptide in vivo to the ascending limb of Henle's loop and the early distal tubules of the kidneys to produce and secrete the heterologous polypeptide into the urine of the transgenic non-human mammal as a recombinant biologically active polypeptide;
- collecting urine from the transgenic non-human mammal;
- and
- recovering the secreted polypeptide as a recombinant biologically active polypeptide.
A transgenic non-human mammal all of whose germ cells and somatic cells contain a recombinant construct corresponding to a DNA molecule comprising a mammalian uromodulin, selected from the group consisting of mouse, goat, human, rat and bovine uromodulin promoters and operably linked to a heterologous DNA sequence encoding a heterologous polypeptide, said DNA molecule having been introduced into said mammal selected from the group consisting of goat, sheep, cattle, pig and mouse at an embryonic stage, wherein said mammalian uromodulin promoter directs expression of said heterologous polypeptide in vivo to the ascending limb of Henle's loop and the early distal tubules of the kidneys to produce recoverable amounts of a recombinant biologically active polypeptide in its urine.
Dependencies are inferred from the claim text.