IP Library Granted Patent US 8,628,957
Granted Patent B2
US 8,628,957 · App. 13/370,795 · Granted Jan 14, 2014

Rat embryonic stem cell

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Quick Facts
Patent No.
US 8,628,957
App. No.
13/370,795
Granted
Jan 14, 2014
Kind
B2
Abstract

The present invention provides a rat embryonic stem cell characterized by having the following properties of (a) expressing Oct3/4 gene and Nanog gene, (b) positive for alkaline phosphatase activity, (c) having an embryoid body forming ability, (d) expressing SSEA (Stage-Specific Embryonic Antigen)-1 and SSEA-4, (e) having the same number of chromosomes as does a normal rat cell, (f) capable of being subcultured and holding the undifferentiated state, (g) having in vitro pluripotency, (h) having a potential to differentiate for cells of three embryonic germ lineages, (i) having teratoma formation ability, and (j) having an ability to produce a chimeric rat, a method of establishing the aforementioned rat embryonic stem cell and the like.

Claims (34)

1. An isolated rat embryonic stem cell characterized by having the following properties (a)-(j):

(a) expressing Oct3/4 gene and Nanog gene,

(b) positive for alkaline phosphatase activity,

(c) having an embryoid body forming ability,

(d) expressing SSEA (Stage-Specific Embryonic Antigen)-1 and SSEA-4,

(e) having the same number of chromosomes as does a normal rat cell,

(f) capable of being subcultured and holding the undifferentiated state,

(g) having in vitro pluripotency,

(h) having a potential to differentiate for cells of three embryonic germ lineages,

(i) having teratoma formation ability,

(j) having an ability to produce a chimeric rat.

2. A rat embryonic stem cell obtained by performing a process consisting essentially of the following steps (A)-(E) using a culture medium with 2% or less serum concentration:

(A) culturing a rat blastocyst on inactivated mouse embryonic fibroblasts in a leukemia inhibitory factor (LIF)-free culture medium to form an inner cell mass in the blastocyst,

(B) dissociating the inner cell mass, wherein the dissociated inner cell mass is in a cell aggregate state,

(C) culturing primary embryonic stem cells resulting from a culture of the dissociated inner cell mass on inactivated mouse embryonic fibroblasts until the primary embryonic stem cells can be passaged,

(D) dissociating the primary embryonic stem cells, which can be passaged, wherein the dissociated primary embryonic stem cells are in a cell aggregate state, and

(E) culturing the dissociated primary embryonic stem cells on inactivated mouse embryonic fibroblasts to establish an embryonic stem cell,

wherein a rat leukemia inhibitory factor (rLIF)-containing culture medium is used in steps (C)-(E).

3. The embryonic stem cell of claim 2 , wherein the culture medium comprises a serum replacement reagent.

4. The embryonic stem cell of claim 3 , wherein step (B) comprises a step for mechanically dissociating the inner cell mass.

5. The embryonic stem cell of claim 4 , wherein step (D) comprises a step for mechanically dissociating the embryonic stem cells.

6. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 1 .

7. The embryonic stem cell of claim 2 , wherein step (B) comprises a step for mechanically dissociating the inner cell mass.

8. The embryonic stem cell of claim 7 , wherein step (D) comprises a step for mechanically dissociating the embryonic stem cells.

9. The embryonic stem cell of claim 2 , wherein step (D) comprises a step for mechanically dissociating the embryonic stem cells.

10. The embryonic stem cell of claim 3 , wherein step (D) comprises a step for mechanically dissociating the embryonic stem cells.

11. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 2 .

12. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 3 .

13. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 4 .

14. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 5 .

15. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 7 .

16. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 8 .

17. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 9 .

18. A culture kit comprising, as components, a serum replacement reagent, rLIF, and the rat embryonic stem cell of claim 10 .

Assignments (4)
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Mar 28, 2019
From: DS PHARMA BIOMEDICAL CO., LTD.
To: SUMITOMO CHEMICAL COMPANY, LIMITED
Reel/Frame 048724/0360 →
ADDRESS CHANGE Recorded Nov 28, 2017
From: DS PHARMA BIOMEDICAL CO., LTD.
To: DS PHARMA BIOMEDICAL CO., LTD.
Reel/Frame 044514/0891 →
ASSIGNMENT OF ASSIGNOR'S INTEREST Recorded Sep 15, 2016
From: SUMITOMO DAINIPPON PHARMA CO., LTD.
To: DS PHARMA BIOMEDICAL CO., LTD.
Reel/Frame 039759/0163 →
CHANGE OF NAME Recorded Aug 25, 2014
From: DAINIPPON SUMITOMO PHARMA CO., LTD.
To: SUMITOMO DAINIPPON PHARMA CO., LTD.
Reel/Frame 033606/0307 →