DNA transformation of a pluripotent mouse embryonal stem cell line with a dominant selective marker

The mouse embryonal stem cell line BLC 1 growing on feeder layer was treated with a calcium phosphate/DNA precipitate prepared with DNA of plasmid pAG60 which harbors the Tn5-derived neo gene, thus encoding resistance to G418, an aminoglycoside antibiotic. Transfection performed on feeder layer resulted in the formation of G418-resistant clones T1 and T2/K26. The stable integration of the transformed neo gene was confirmed by dot hybridization in all descendant cultures of clones T1 and T2/K26 as well as in the tumors derived from them. In vivo and in vitro differentiation revealed the pluripotent status of the transformants. Tumors derived from T1 and T2/K26 contained various tissues with derivatives of all three primary germ layers.