ISSN:
1573-0778
Keywords:
beta-interferon
;
dihydrofolate reductase
;
gene coamplification
;
Namalwa KJM-1
;
serum-free medium
Source:
Springer Online Journal Archives 1860-2000
Topics:
Biology
,
Medicine
,
Process Engineering, Biotechnology, Nutrition Technology
Notes:
Abstract We previously reported the expression of human beta-interferon (β-IFN) (Miyajiet al., 1989) and human lymphotoxin (Miyajiet 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 β-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 β-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 β-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 β-IFN at a level as high as 5 μg/ml, which is about 100-fold higher than that obtained by the G418-resistant parental transformants. In addition, β-IFN produced by recombinant KJM-1 cells had the same molecular weight of that produced by fibroblasts.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00365098
