Cultured inner cell mass cell lines derived from bovine or porcine embryos

Claim 1

A method for producing a bovine or porcine cultured inner cell mass (CICM) cell which may be incorporated into a bovine or porcine embryo to form a chimeric bovine or porcine which consists essentially of the following steps: (i) obtaining inner cell mass (ICM) cells from a bovine or porcine blastocyst or ICM progenitor cells from a bovine or porcine preblastocyst stage embryo; (ii) culturing said ICM or ICM progenitor cells on a feeder layer culture to produce multilayer cell colonies; (iii) identifying from among the cells contained in the cultured ICM or ICM progenitor cell colony those cells which exhibit the following properties: (a) small cytoplasmic/nuclear volume ratio ranging from about 10/90 to 50/50; (b) cytoplasmic vesicles; (c) small individual cells ranging from about 10 to 20 μm in diameter; (iv) separating one or a cluster of said identified cells from the rest of the cell colony; and (v) passaging said separated ICM or ICM progenitor cells onto another feeder layer culture and maintaining physical contact between the feeder cell layer and the separated cell or cell cluster throughout the entire culturing period to produce CICM cells that may be incorporated into a bovine or porcine embryo to form a chimeric bovine or porcine.

Claim 2

A method for producing a bovine or porcine cultured inner cell mass (CICM) cell which may be incorporated into a bovine or porcine embryo to form a chimeric bovine or porcine which consists essentially of the following steps: (i) obtaining inner cell mass (ICM) cells from a bovine or porcine blastocyst or ICM progenitor cells from a bovine or porcine preblastocyst stage embryo; (ii) culturing said ICM or ICM progenitor cells on a feeder layer culture to produce multilayer cell colonies; (iii) identifying from among the cells contained in the cultured ICM or ICM progenitor cell colony those cells which exhibit the following properties: (a) small cytoplasmic/nuclear volume ratio ranging from about 10/90 to 50/50; (b) cytoplasmic vesicles; (c) small individual cells ranging from about 10 to 20 μm in diameter; (iv) separating one or a cluster of said identified cells from the rest of the cell colony; and (v) passaging said separated ICM or ICM progenitor cells onto another feeder layer culture and maintaining physical contact between the feeder cell layer and the separated cell or cell cluster throughout the entire culturing period to produce CICM cells that may be incorporated into a bovine or porcine embryo to form a chimeric bovine or porcine.