稳定转化家蚕BmN细胞表达人粒细胞-巨噬细胞集落刺激因子  

Expression of hGM-CSF in Stably Transformed BmN Cells of Bombyx mori

作　　者：李曦[1] 赵越[1] 周文林[1] 曹广力[1] 薛仁宇[1] 贡成良[1] 

机构地区：[1]苏州大学基础医学与生物科学学院,江苏苏州215123

出　　处：《蚕业科学》2009年第2期302-307,共6页ACTA SERICOLOGICA SINICA

基　　金：国家重点基础研究发展计划"973"项目(编号2005CB12-1000)

摘　　要：为了建立稳定转化家蚕细胞持续表达外源基因的技术体系,构建了基于piggyBac转座子的带有人粒细胞-巨噬细胞集落刺激因子基因(hGM-CSF)和新霉素抗性基因(neo)表达元件的转基因载体pigA3GFP-IE-neo-FH-hGM-CSF-polyA-fib-L-intron1,以及带有hGM-CSF和吉欧霉素(zeocin)抗性基因的昆虫细胞转化载体pIZT-IE-hGM-CSF,分别转染家蚕卵巢BmN细胞,并以含相应抗生素G418或zeocin的培养液筛选,得到稳定的转化细胞系FH-hGM-CSF和IE-hGM-CSF。ELISA检测结果显示,hGM-CSF在FH-hGM-CSF和IE-hGM-CSF转化细胞系的表达水平分别为1.534 55 fg/个细胞和2.227 38 fg/个细胞。In order to generate stable transformed Bombyx mori cell line for expressing target gene continuously, two transgenic vectors, pigA3GFP-IE-neo-FH-hGM-CSF-polyA-fib-L-intronl containing a human granulocyte-macrophage colony-stimulating factor （hGM-CSF） gene and a neomycin resistance gene based on piggyBac transposon, and plZT-IE-hGM-CSF containing a hGM-CSF gene and a zeocin resistance gene were constructed. Silkworm ovarian Bran cells were transfected with the two transgenic vectors respectively, and screened with antibiotics G418 or zeocin. Two stable transformed cell lines named FH-hGM-CSF and IE-hGM-CSF were obtained respectively. ELISA assay indicated that the expression level of hGM-CSF in cell lines FH- hGM-CSF and IE-hGM-CSF was about 1. 534 55 fg and 2. 227 38 fg percell, respectively.

关 键 词：PIGGYBAC转座子 pIZT/V5-His载体 家蚕BmN细胞 人粒细胞-巨噬细胞集落刺激因子 

分 类 号：S881.2[农业科学—特种经济动物饲养] Q813[农业科学—畜牧兽医]