Expression of dominant negative transmembrane receptors in the milk of transgenic animals
First Claim
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1. A method for cloning a non-human mammal through a nuclear transfer process comprising:
- (i) obtaining desired differentiated mammalian cells to be used as a source of donor nuclei;
(ii) obtaining at least one oocyte from a mammal of the same species as the cells which are the source of donor nuclei;
(iii) enucleating said at least one oocyte;
(iv) transferring the desired differentiated cell or cell nucleus into the enucleated oocyte;
(v) simultaneously fusing and activating the cell couplet to form a transgenic embryo;
(vii) culturing said transgenic embryo(es) until greater than the 2-cell developmental stage; and
(viii) transferring said transgenic embryo into a host mammal such that the embryo develops into a fetus;
wherein the desired differentiated cell or cell nucleus contains a recombinant transgene; and
, wherein said recombinant transgene encodes a recombinant transmembrane receptor protein of interest.
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Abstract
The present invention provides data to demonstrates the transgenic mammal production of membrane spanning receptor proteins in the milk of transgenic animals, offering a method of production of these proteins and dominant negative versions thereof for use as therapeutic molecules.
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Citations
75 Claims
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1. A method for cloning a non-human mammal through a nuclear transfer process comprising:
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(i) obtaining desired differentiated mammalian cells to be used as a source of donor nuclei;
(ii) obtaining at least one oocyte from a mammal of the same species as the cells which are the source of donor nuclei;
(iii) enucleating said at least one oocyte;
(iv) transferring the desired differentiated cell or cell nucleus into the enucleated oocyte;
(v) simultaneously fusing and activating the cell couplet to form a transgenic embryo;
(vii) culturing said transgenic embryo(es) until greater than the 2-cell developmental stage; and
(viii) transferring said transgenic embryo into a host mammal such that the embryo develops into a fetus;
wherein the desired differentiated cell or cell nucleus contains a recombinant transgene; and
,wherein said recombinant transgene encodes a recombinant transmembrane receptor protein of interest. - View Dependent Claims (2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 48, 49, 50, 51, 52, 53, 54, 55, 56)
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24. A method for producing cultured inner cell mass cells, comprising:
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(i) obtaining desired differentiated mammalian cells to be used as a source of donor nuclei;
(ii) obtaining at least one oocyte from a mammal of the same species as the cells which are the source of donor nuclei;
(iii) enucleating said at least one oocyte;
(iv) transferring the desired differentiated cell or cell nucleus into the enucleated oocyte;
(v) simultaneously fusing and activating the cell couplet to form a first transgenic embryo;
(vi) activating a cell-couplet that does not fuse to create a first transgenic embryo but that is activated after an initial electrical shock by providing at least one additional activation protocol including an additional electrical shock to form a second transgenic embryo; and
(vi) culturing cells obtained from said cultured activated embryo to obtain cultured inner cell mass cells;
wherein said transgenic embryo encodes a recombinant transmembrane receptor protein of interest. - View Dependent Claims (25, 26, 27, 28, 29, 30, 31, 32, 33, 34, 35, 36, 37, 38, 39, 40, 41, 42, 43, 44, 45, 46, 47, 57, 58, 59, 60, 61, 62, 63, 64, 65)
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66. A method for cloning a non-human mammal through a nuclear transfer process comprising:
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(i) obtaining desired differentiated mammalian cells to be used as a source of donor nuclei;
(ii) obtaining at least one oocyte from a mammal of the same species as the cells which are the source of donor nuclei;
(iii) enucleating said oocytes;
(iv) transferring the desired differentiated cell or cell nucleus into the enucleated oocyte;
employing at least two electrical shocks to a cell-couplet to initiate fusion and activation of said cell-couplet into an activated and fused embryo. (vii) culturing said activated and fused embryo until greater than the 2-cell developmental stage;
(viii) transferring said fused embryo into a host mammal such that the embryo develops into a fetus;
wherein the second of said at least two electrical shocks is administered at least 15 minutes after an initial electrical shock;
wherein a desired gene is inserted, removed or modified in said differentiated mammalian cell or cell nucleus prior to insertion of said differentiated mammalian cell or cell nucleus into said enucleated oocyte; and
wherein said desired gene encodes a recombinant transmembrane receptor protein of interest that can be expressed upon induction of lactation in mammary epithelial cells.
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- 67. A method of treating a disease comprising the administering of an effective amount of a transgenically produced transmembrane receptor protein or dominant negative version thereof such that said compound comes into contact with a cell or group of cells which have been or will be exposed to a disease condition where said compound acts to interfere with the continued progression of the disease.
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73. A method of treating obesity comprising the administering of an effective amount of a transgenically produced transmembrane receptor protein or dominant negative version thereof.
Specification