| 磷酸钙法对哺乳动物非贴壁细胞的质粒共转染 |
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| 引用本文： | 李水仙,赵连三,贾文祥,唐红,刘丽,刘聪,丁柳,王松.磷酸钙法对哺乳动物非贴壁细胞的质粒共转染[J].生物技术通讯,1998(1). |
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| 作者姓名： | 李水仙 赵连三 贾文祥 唐红 刘丽 刘聪 丁柳 王松 |
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| 作者单位： | 华西医科大学附一医院肝炎研究室，华西医科大学附一医院肝炎研究室，华西医科大学附一医院微生物教研室，华西医科大学附一医院肝炎研究室，华西医科大学附一医院肝炎研究室，华西医科大学附一医院肝炎研究室，华西医科大学附一医院肝炎研究室，华西医科大学附一医院肝炎研究室 研究生，山西长治医学院微生物教研室 长治 046000，成都 610041 |
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| 摘 要： | 对一种哺乳动物的非贴壁细胞Bal b/c近交系小鼠肥大细胞瘤细胞株P815进行了G418敏感浓度及甘油耐受性的测定;并通过磷酸钙法用含HBVS基因的质粒pRc/CMVS与含筛选标记基因neo的质粒pSV_2-neo进行了共转染。经G418筛选,获得了G418抗性细胞P851-S,在G418选择压力下,已连续筛选80余天,传代12次;斑点杂交和ELISA实验分别表明P815-S细胞内有HBVS基因的存在,培养上清中有HBsAg的表达。
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| 关 键 词： | 共转染 质粒 表达 非贴壁细胞 甘油“休克” |
Plasmids cotransfections to non-attaching-wall mammalian cells using calcium phosphate method |
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| Li Shuixian,Zhao Liansan,Jia Wenxiang,Tang Hong,Liu Li,Liu Cong,Ding Liu,Wang Song.Plasmids cotransfections to non-attaching-wall mammalian cells using calcium phosphate method[J].Letters in Biotechnology,1998(1). |
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| Authors: | Li Shuixian Zhao Liansan Jia Wenxiang Tang Hong Liu Li Liu Cong Ding Liu Wang Song |
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| Abstract: | Bal b/c inbred mice mastocytoma cell line P815 (a kind of non-attaching-wall mammalian cells) was tested to determine its sensitive concentration to G418 and whether it could tolerate glycerine; then, with calcium phosphate method, P815 cells were cotransfected by a plasmid (pRc/CMVS) containing HBV-S gene and a plasmid (pSV2-neo) containing neomycin-recistance gene (marker gene). By selection with G418, G418 resistant cells (P815-S) were obtained, and were already selected under the pressure of G418 for more than 80 days, 12 generations continuously. The results of dot-blot and hybnding and ELISA showed that HBV-S genes exist in P815-S cells; HBVS proteins are secreted into culture medium of P815-S cells. These results indicated that cotransfection was successful. |
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| Keywords: | cotransfection plasmid expression non-attaching-well cells glycerine "shock |
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