Genetic modification of rats

An isolated population of rat ES cells, wherein at least 70% of the rat ES cells are euploid and form sphere-like colonies when plated on a feeder cell layer in vitro.

The targeted genetic modification comprises at least one insertion of a heterologous polynucleotide into a genome of the cell.

An in vitro culture comprising a feeder cell layer, the population of rat embryonic stem (ES) cells, and a medium comprising a Leukemia Inhibitory Factor (LIF), GSK3 inhibitor, and a MEK inhibitor, wherein at least 70% of the rat ES cells are euploid and the rat ES cell forms a sphere-like colony.

A method for generating a rat embryonic stem (ES) cell line comprising:

• (a) culturing in vitro a first feeder cell layer and a morula or a blastocyst-stage rat embryo, wherein the zona pellucida of the morula or blastocyst-stage rat embryo has been removed, and wherein the culture conditions maintain pluripotency of a rat ES cell and comprise a medium having mouse leukemia inhibitor factor (LIF) or a sequence having at least 91% sequence identity to SEQ ID NO:1 and having LIF activity, and a GSK3 inhibitor, and a MEK inhibitor;
• and, (b) transferring an outgrowth of an amorphous undifferentiated mass of rat ES cells to an in vitro culture well comprising a second feeder cell layer and culturing the outgrowth under conditions comprising the medium having the mouse LIF or an active variant of the mouse LIF, and thereby maintaining pluripotency of the rat ES cells;
• and, establishing a rat ES cell line therefrom.