Miyaji H, Mizukami T, Hosoi S, Sato S, Fujiyoshi N, Itoh S
Tokyo Research Laboratories, Kyowa Hakko Kogyo Co., Ltd., Japan.
Cytotechnology. 1990 Mar;3(2):133-40. doi: 10.1007/BF00143675.
A Namalwa cell line, KJM-1, which was adapted to serum-free medium is thought to be a very useful host cell line for recombinant DNA technology. Thus, the utility of Namalwa KJM-1 for expression of foreign genes was examined. As a model system human beta-interferon (beta-IFN) gene was engineered for expression in this cell line. For construction of the beta-IFN expression vector pSE1 beta 1-4, the expression vector pAGE107 was constructed and used. It contains simian virus 40 (SV40) early promoter, the rabbit beta-globin RNA processing signals for splicing and polyadenylation, and SV40 early RNA processing signal for polyadenylation. In addition to the above transcription unit, pAGE107 contains the ampicillin-resistance gene and G418-resistance gene. They can confer ampicillin resistance to Escherichia coli (E. coli) and G418 resistance to animal cells. To introduce plasmid DNA into cells, electroporation is a useful method (Wong, 1982; Potter, 1984). We have established conditions for DNA-mediated transfection of Namalwa KJM-1 cell line by electroporation. Among pSE1 beta 1-4-introduced cells, clone 1-3 was further examined for the expression of beta-IFN in serum-free medium. The production level of beta-IFN was elevated with the increase of the cell density. The results indicated that the Namalwa KJM-1 cell line is useful for production of foreign gene products.
一种适应无血清培养基的Namalwa细胞系KJM - 1，被认为是重组DNA技术中非常有用的宿主细胞系。因此，研究了Namalwa KJM - 1在表达外源基因方面的实用性。作为一个模型系统，构建了人β - 干扰素（β - IFN）基因以便在该细胞系中表达。为构建β - IFN表达载体pSE1β1 - 4，构建并使用了表达载体pAGE107。它包含猿猴病毒40（SV40）早期启动子、用于剪接和聚腺苷酸化的兔β - 珠蛋白RNA加工信号以及用于聚腺苷酸化的SV40早期RNA加工信号。除上述转录单元外，pAGE107还包含氨苄青霉素抗性基因和G418抗性基因。它们可使大肠杆菌（E. coli）获得氨苄青霉素抗性，使动物细胞获得G418抗性。为了将质粒DNA导入细胞，电穿孔是一种有用的方法（Wong，1982；Potter，1984）。我们已经建立了通过电穿孔对Namalwa KJM - 1细胞系进行DNA介导转染的条件。在导入pSE1β1 - 4的细胞中，对克隆1 - 3在无血清培养基中β - IFN的表达进行了进一步检测。β - IFN的产生水平随着细胞密度的增加而升高。结果表明Namalwa KJM - 1细胞系可用于生产外源基因产物。
