rhGM-CSF在真核细胞中的长期稳定表达及糖基化  

Long-term stable expression and glycosylation of rhGM-CSF in eukaryotic cells

作　　者：戴萍[1] 魏大鹏[1] 章崇杰[1] 蔡美英[1] 

机构地区：[1]华西医科大学免疫教研室,四川成都610041

出　　处：《川北医学院学报》2001年第2期3-6,共4页Journal of North Sichuan Medical College

基　　金：四川省科委应用基础研究专项基金资助

摘　　要：通过重组DNA技术 ,经RT -PCR从人外周血淋巴细胞中获得全长cDNA序列 ,连入pcDNA3 .1(+ )中 ,构建了可以在哺乳动物细胞稳定表达的质粒pcDNA -hGM -CSF ,并用脂质体法转入CHO细胞 ,经G4 18筛选 ,建立起长期稳定分泌hGM -CSF的细胞株 ,在三个月的观察中分泌量恒定在 43.49± 8.45 6ng/ml,活性单位达 480IU/ml/ 10 5个细胞 ,Western -blot证实该细胞株分泌的hGM -CSF有 6种分子量 ,主带在 17KD及 35KD左右 ,提示其有不同程度糖基化。By RT-PCR,the full-length cDNA of rhGM-CSF was amplified from the peripheral blood lymphocytes of healthy adult,and then cloned into the plasmid pcDNA33.1(+).After recombined vector pcDNA3-rhGM-CSF transfected the CHO cells,we got the drug-resistant CHO in G 418 medium.It can stably secrete rhGM-CSF at least for three months,the average product is 43.49±8.456 ng/ml,and the activity is about 480IU/ml/105 cells.By the Western-blot,we find that hGM-CSF has six MW in supernatant, mainly 17KD and 35KD, this result demonstrated the rhGM-CSF can be glycosylated by CHO.

关 键 词：重组hGM-CSF RT-PCR 真核表达 糖基化 真核细胞 

分 类 号：R392.12[医药卫生—免疫学]