Method of Differentiation of Morula or Inner Cell Mass Cells and Method of Making Lineage-Defective Embryonic Stem Cells

Abstract

An improved method of producing differentiated progenitor cells comprising obtaining inner cell mass cells from a blastocyst and inducing differentiation of the inner cell mass cells to produce differentiated progenitor cells. The differentiated progenitor cells may be transfected such that there is an addition, deletion or alteration of a desired gene. The differentiated progenitor cells are useful in cell therapy and as a I source of cells for the production of tissues and organs for transplantation. Also provided is a method of producing a lineage-defective human embryonic stem cell.

28 . A method of producing a lineage-defective embryonic stem cell, comprising:
 (I) genetically modifying a somatic cell such that said somatic cell is incapable of differentiating into a predetermined cell lineage;   (II) generating a nuclear transfer unit using the genetically modified somatic cell or cell nucleus as the nuclear donor;   (III) activating the resultant nuclear transfer unit;   (IV) culturing said activated nuclear transfer unit until greater than the 2-cell developmental stage; and   (V) culturing cells obtained from said cultured nuclear transfer unit under conditions suitable for the formation of a lineage-defective embryonic stem cell,   
       said stem cell being unable to differentiate into at least one of the embryonic germ layers. 
     
     
         29 . The method according to  claim 29 , wherein generating said nuclear transfer unit comprises inserting the genetically modified human somatic cell or cell nucleus into an enucleated mammalian oocyte under conditions suitable for formation of a nuclear transfer unit. 
     
     
         30 . The method according to  claim 29 , wherein said lineage-defective human embryonic stem cell is incapable of differentiating into mesoderm. 
     
     
         31 . The method according to  claim 29 , wherein said lineage-defective human embryonic stem cell is incapable of differentiating into endoderm. 
     
     
         32 . The method according to  claim 29 ; wherein said lineage-defective human embryonic stem cell is incapable of differentiating into ectoderm. 
     
     
         33 . A lineage-defective human embryonic stem cell produced according to the method of  claim 29 . 
     
     
         34 . The method according to  claim 29 , wherein said lineage-defective embryonic stem cell is human. 
     
     
         36 . The method of  claim 28  wherein the somatic cell comprises a genetic construct comprising an inducible promoter operably linked to a gene the expression of which blocks the growth of undifferentiated cells. 
     
     
         37 . The method of  claim 28 , wherein the somatic cell comprises a genetic construct that comprises a promoter that is germ-line specific that regulates the expression of cell cycle blocker or an apoptosis gene. 
     
     
         38 . The method of  claim 28 , wherein the somatic cell comprises a genetic construct that comprises an inducible promoter operably linked to a gene that induces the differentiation of undifferentiated cells. 
     
     
         39 . The method of  claim 36 , wherein the somatic cell is human. 
     
     
         40 . The method of  claim 37 , wherein the somatic cell is human. 
     
     
         41 . The method of  claim 38 , wherein the somatic cell is human. 
     
     
         42 . An isolated embryonic cell that comprises an inducible promoter operably linked to a gene the expression of which prevents the growth of undifferentiated cells. 
     
     
         43 . The embryonic cell of  claim 42  which is human. 
     
     
         45 . The embryonic cell of  claim 43  which is human. 
     
     
         46 . The embryonic cell of  claim 42  which is derived from a nuclear transfer embryo. 
     
     
         47 . The embryonic cell of  claim 43  which is from a nuclear transfer embryo. 
     
     
         48 . The embryonic cell of  claim 42  which is an inner cell mass cell or a morula cell. 
     
     
         49 . The embryonic cell of  claim 43  which is an inner cell mass or morula cell.