Glutamine synthetase gene amplification system was applied to develop erytoropoeitin(EPO) producing cell line. EPO cDNA obtained by PCR was cloned into expression vector CDM8-GS haboring glutamine synthetase(GS) gene as a selection marker, and this construct was transfected into Chinese Hamster Ovary(CHO) cells using various transfection methods. Transfectants were selected in glutamine free media containing 20μM methionine sulfoximine(MSX) for about 2 weeks. However, stable transfectants were not selected, so that several possible and reasonable factors such as MSX concentration, suitability of glutamine free media, transfection efficiency were checked but any problems were not found. To overcome this problem, an alternative method of selection, sequential selection, was designed. Neo marker gene that has resistance to Geneticin(G-418) was cloned into GS-EPOc expression vector, and transfectants were selected with DMEM containing G-418. Stable transfectants were obtained through selection procedure, and chromosomal integration of vector was confirmed by the detection of EPO with ELISA. And then these cells were cultured in the selection media of GS marker as a second selection, but no stable transfectants was emerged after selection. Meanwhile, the mechanism of cell death during GS marker selection was investigated. As a result, apoptosis was observed and confirmed by the detection of chromosomal DNA fragmentation. The reason of apoptosis was supposed to be glutamine deficiency in selection media.