将人白细胞介素12(interleukin 12, IL-12)两条链p35及p40全长cDNA分别亚克隆至真核表达载体pcDNA3中,构建了pcDNA3/p35a,pcDNA3/p40a,pcDNA3/p35b,pcDNA3/p40b四种真核细胞重组表达质粒,利用磷酸钙共沉淀法转染中国苍鼠卵巢(CHO)细胞. 通过对阳性克隆的筛选鉴定,获得了稳定表达人IL-12的CHO细胞株,活性最高的一株表达量为105 U/ml.此细胞株经半年的传代培养,能够稳定地分泌IL-12.结果显示:IL-12在CHO细胞中的稳定表达受到重组质粒结构、DNA整合、mRNA转录、蛋白质翻译等多因素的影响,IL-12两个亚基在CHO细胞中的共同表达是产生有生物活性IL-12的基础.
Four kinds of human IL-12 expression plasmids pcDNA3/p35a, pcDNA3/p35b, pcDNA3/p40a, pcDNA3/p40b were constructed. These recombinant plasmids were co-transfected into Chinese hamster ovary cells and acquired CHO cell lines that could stably secrete IL-12. The highest expressed amount of IL-12 in CHO cell is 105 U/ml. The results showed that the expression of human IL-12 in CHO cell was affected at several levels such as recombinant plasmid structure, DNA intergration, mRNA transcription and protein transpation etc.
戴燕,陈慰峰.人重组白细胞介素12在CHO细胞中的表达[J].生物化学与生物物理进展,1998,25(3):254-258
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