Efficient expression of human beta-interferon in Namalwa KJM-1 cells adapted to serum-free medium by a dhfr gene coamplification method.
We previously reported the expression of human beta-interferon (beta-IFN) (Miyaji et al., 1989) and human lymphotoxin (Miyaji et al., 1990) in Namalwa KJM-1 cells adapted to serum-free medium. To establish an efficient gene expression system, a dihydrofolate reductase (dhfr) gene coamplification method was applied to this cell line. A beta-IFN expression plasmid was introduced with a dhfr expression plasmid into KJM-1 and methotrexate (MTX)-resistant derivatives were selected by a stepwise increase of MTX concentration. Among them, derivatives which showed higher expression levels of beta-IFN than that achieved by the parental transformants were obtained, suggesting that a dhfr gene coamplification method can be used for efficient expression of foreign genes in KJM-1 which contains endogenous dhfr genes. Then, an improved beta-IFN expression vector was constructed, which contains a dhfr transcription unit. This plasmid was introduced into KJM-1 and then, MTX-resistant derivatives were selected. Among them, the highest producer, clone 40-10-24, secreted beta-IFN at a level as high as 5 micrograms/ml, which is about 100-fold higher than that obtained by the G418-resistant parental transformants. In addition, beta-IFN produced by recombinant KJM-1 cells had the same molecular weight of that produced by fibroblasts.
Miyaji
H
H
Tokyo Research Laboratories, Kyowa Hakko Kogyo Co., Ltd., Japan.
Harada
N
N
Mizukami
T
T
Sato
S
S
Fujiyoshi
N
N
Itoh
S
S
D015153:Blotting, Western; D002460:Cell Line; D003470:Culture Media; D005784:Gene Amplification; D005822:Genetic Vectors; D006801:Humans; D007370:Interferon Type I / Q000096:biosynthesis* / Q000235:genetics; D008214:Lymphocytes / Q000378:metabolism; D011994:Recombinant Proteins; D013762:Tetrahydrofolate Dehydrogenase / Q000235:genetics*; D014162:Transfection
D003470:Culture Media; D007370:Interferon Type I; D011994:Recombinant Proteins; D013762:Tetrahydrofolate Dehydrogenase