Compositions for the derivation of germ cells from stem cells and methods of use thereof

1. A method for deriving oocytes from mouse embryonic stem (ES) cells comprising:

(a) transforming mouse embryonic stem cells with a germ cell specific promoter sequence operably linked to a reporter gene;

(b) culturing the transformed cells in ES cell medium under conditions which promote differentiation;

(c) optionally removing the differentiated cells from step b) from said conditions which promote differentiation;

(d) monitoring the cells of step (b) or (c) for reporter gene expression thereby identifying germ cells in a mixed cell population; and

(e) isolating oocytes from the germ cells identified from step (d);

wherein said reporter gene encodes a fluorescent protein and

wherein said germ cell specific promoter sequence is a truncated Oct4 promoter.

2. The method of claim 1 , wherein said oocytes are isolated via a method selected from the group consisting of flow cytometry, enzymatic digestion, and microdissection.

3. The method of claim 1 , wherein said differentiation in step b) is promoted by culturing said cells in the absence of leukemia inhibitory factor (LIE).

4. The method of claim 1 , wherein said removing of cultured cells from said conditions which promote differentiation in step (c) comprises replating said cultured cells in media comprising LIE, basic fibroblast growth factor (b-EGF) and stem cell factor.

5. The method of claim 1 , optionally further comprising subjecting said germ cells identified in step d) to culture conditions which promote the formation of oocyte structures.

6. The method of claim 5 , wherein said oocyte structures express zona pellucida protein 3 (ZP3).

7. The method of claim 1 , wherein said reporter gene is green fluorescent protein.

8. The method of claim 1 , wherein said truncated Oct4 promoter gene sequence is an Oct4 promoter gene sequence with conserved regions 2 and 3 deleted.

9. The method of claim 8 , wherein said truncated Oct4 promoter gene sequence is SEQ ID NO: 1.

10. An oocyte obtained by the method of claim 1 , wherein said truncated Oct4 promoter gene sequence is an Oct4 promoter gene sequence which lacks conserved regions 2 and 3.

11. An oocyte obtained via the method of claim 5 , wherein said truncated Oct4 promoter gene sequence is an Oct4 promoter gene sequence which lacks conserved regions 2 and 3.

12. A method for deriving oocytes from mouse embryonic stem (ES) cells comprising:

(a) culturing the mouse embryonic stem cells in ES cell medium under conditions which promote differentiation;

(b) optionally removing the differentiated cells from step a) from said conditions which promote differentiation;

(c) monitoring the cells of step (a) or (b) for morphological alterations associated with germ cell formation; and

(d) isolating oocytes from the germ cells identified from the morphological alterations observed in step (c).

13. The method of claim 12 , wherein said morphological alterations comprise at least one of the following: cell enlargement, increased spherical appearance, and reduced cell to cell contacts.

14. The method of claim 12 , wherein said oocytes are isolated via a method selected from the group consisting of flow cytometry, enzymatic digestion, and microdissection.

15. The method of claim 12 , wherein said differentiation in step a) is promoted by culturing said cells in the absence of leukemia inhibitory factor (LIE).

16. The method of claim 12 , wherein said removing of cultured cells from said conditions which promote differentiation in step (b) comprises replating said cultured cells in media comprising LIE, basic fibroblast growth factor (b-FGF) and stem cell factor.

17. The method of claim 12 , optionally further comprising subjecting said germ cells identified in step c) to culture conditions which promote the formation of oocyte structures.

18. The method of claim 17 , wherein said oocyte structures express zona pellucida protein 3 (ZP3).

19. A method for screening test compounds for toxicity or teratogenic potential comprising:

a) providing an oocyte as claimed in claim 5 ;

b) exposing said oocytes to increasing amounts of said test compound; and

c) culturing said oocytes under conditions that promote blastocyst formation; and

d) determining the effect, if any, of said test compound on the ability of said oocyte to form a blastocyst.