用逆转录病毒载体表达人粒细胞－巨噬细胞集落刺激因子  

USE OF RETROVIRAL VECTOR FOR EXPRESSION OFHUMAN GRANULOCYTE-MACROPHAGE COLONYSTIMULATING FACTOR

作　　者：陈庆华[1] 张智清[1] 宋永胜[1] 李福胜[1] 刘红兵[1] 姚立红[1] 李玉英[1] 侯云德[1] 

机构地区：[1]中国预防医学科学院病毒学研究所,北京病毒基因工程国家重点实验室

出　　处：《病毒学报》1995年第2期138-143,共6页Chinese Journal of Virology

摘　　要：应用基因工程的方法，将含有巨细胞病毒（ＣＭＶ）启动子的基因片段和人粒细胞－巨噬细胞集落刺激因子（ｈＧＭ－ＣＳＦ）的ｃＤＮＡ，克隆进逆转录病毒载体Ｎ２Ａ，得到重组质粒Ｎ２Ａ／ＣＭＶ／ｈＧＭ－ＣＳＦ．经脂质体包装并转染包装细胞，通过Ｇ４１８药物筛选，得到抗性克隆。经ＰＣＲ和Ｓｏｕｔｈｅｍｂｌｏｔ检测证实，ＧＭ－ＣＳＦ基因已整合到该克隆细胞的染色体上，获得的逆转录病毒滴度达１０￣４ＣＦＵ／ｍｌ，克隆细胞培养上清用ＴＦ－１细胞可检测到ＧＭ－ＣＳＦ活性。he gene of human granulocyte-macrophage colony stimulating factor under the controlof CMV promoter was cloned into retroviral vector N2A to construct the expressing vectorN2A/CMV/hGM-CSF.This recombinant polasmid was transfected into packaging cell line bylipofectin and colonies capable of secreting recombinant retrovirus and GM-CSF were selectedby G418.The integration of GM-CSF gene was detected by PCR and Southerm blothybridization.The titer of recombinant retrovirus is about 10￣4CFU/ml.The supernatant of thecolonies can support the growth of TF-1 cells.

关 键 词：重组逆转录病毒 人GM-CSF 基因工程 研制 

分 类 号：R392－33[医药卫生—免疫学] R394.8[医药卫生—基础医学]