Selectable markers and related methods
The invention relates to newly identified selectable marker systems, cells for use in a selectable marker system, and methods for using the selectable marker systems.
1. An auxotrophic mammalian cell genetically engineered to disrupt expression of a protein, said cell comprising:
a gene disruption of a gene encoding a glycine N-methyltransferase,
wherein said glycine N-methyltransferase is inactive or not expressed in said auxotrophic mammalian cell; and
wherein said auxotrophic mammalian cell is unable to grow or undergoes cell death in the absence of cysteine.
2. The auxotrophic mammalian cell of claim 1 ,
wherein said auxotrophic mammalian cell further comprises an exogenous nucleic acid, wherein said exogenous nucleic acid comprises an expressible nucleic acid comprising a nucleic acid encoding a protein corresponding to said inactive or not expressed glycine N-methyltransferase in said auxotrophic mammalian cell.
3. The auxotrophic mammalian cell of claim 2 ,
wherein said exogenous nucleic acid further comprises an expressible nucleic acid encoding a product.
4. A composition comprising a cell, wherein said cell comprises a gene disruption of a glycine N-methyltransferase, wherein said glycine N-methyltransferase is inactive or not expressed in said cell, and said cell further comprises an expressible nucleic acid comprising a nucleic acid encoding a protein corresponding to said inactive or not expressed glycine N-methyltransferase in said cell.
5. The composition of claim 4 , wherein said exogenous nucleic acid further comprises an expressible nucleic acid encoding a product.
6. A kit comprising the composition of claim 4 .
7. The kit of claim 6 , wherein the kit further comprises the components of a selection medium for culturing said cell, wherein the components of the selection medium exclude cysteine.
8. The kit of claim 7 , wherein the kit further comprises cysteine.
9. A method of selecting for an exogenous nucleic acid in a cell, comprising:
(a) contacting a plurality of cells of claim 1 with an exogenous nucleic acid encoding a glycine N-methyltransferase, wherein said contacting results in uptake of said exogenous nucleic acid into at least one of said plurality of cells;
(b) placing said plurality of cells in a selection medium, wherein the components of the selection medium exclude cysteine; and
(c) culturing said plurality of cells in said selection medium, whereby culturing said plurality cells selects for at least one cell that expresses from said exogenous nucleic acid said glycine N-methyltransferase.
10. The method of claim 9 , wherein said exogenous nucleic acid further comprises an expressible nucleic acid encoding a product.
11. The method of claim 9 , wherein said product is expressed in said selected cell.
12. The method of claim 11 , further comprising isolating the product expressed in the selected cell.
13. A method of expressing a product in a cell, comprising:
(a) contacting a plurality of cells of claim 1 with an exogenous nucleic acid encoding glycine N-methyltransferase and an expressible nucleic acid encoding said product, wherein said contacting results in uptake of said exogenous nucleic acid into at least one of said plurality of cells;
(b) placing said plurality of cells in a selection medium, wherein the components of the selection medium exclude cysteine;
(c) culturing said plurality of cells in said selection medium, whereby culturing said plurality of cells selects for at least one cell that expresses from said exogenous nucleic acid said glycine N-methyltransferase; and
(d) culturing said selected cell under conditions for expression of said product.
14. The method of claim 13 , further comprising isolating the product expressed in the selected cell.
15. An auxotrophic mammalian cell comprising:
(a) a gene disruption of a glycine N-methyltransferase,
wherein said glycine N-methyltransferase is inactive or not expressed in said auxotrophic mammalian cell; and
(b) a mammalian expression vector, said mammalian expression vector comprising an expressible nucleic acid comprising a nucleic acid encoding a protein corresponding to said inactive or not expressed glycine N-methyltransferase.
16. The auxotrophic mammalian cell of claim 15 , wherein said auxotrophic mammalian cell is unable to grow or undergoes cell death in the absence of cysteine.
17. The auxotrophic mammalian cell of claim 15 , wherein said exogenous nucleic acid further comprises an expressible nucleic acid encoding a product.