癌胚抗原与IL-2真核双表达质粒的构建及体外表达

Construction of mammalian co-expression plasmid pIRES-CEA-IL-2 and its expression in vitro

目的 :构建癌胚抗原 (CEA)与白细胞介素 2 (IL 2 )的真核双表达质粒 ,并检测其在体外的表达。方法 :利用分子生物学技术 ,将CEA基因片段和IL 2基因片段连接于真核双表达质粒 pIRES中 ,测定序列后 ,用脂质体包裹转染细胞 ,采用ELISA双抗体夹心法检测CEA和IL 2的表达。结果 :核酸序列测定证实本实验所构建的质粒正确 ,质粒上所接入的CEA和IL 2的碱基序列与标准序列的同源性分别为 99.8%和 99.9%。该双表达质粒在体外转染细胞内后可表达CEA和IL 2分子。结论 :实验所构建的 pIRES CEA IL 2双表达质粒能在体外同时表达CEA和IL 2分子 ,说明本质粒具有在细胞水平同时表达两种生物大分子的活性 。

Purpose:To construct the mammalian co expression plasmid pIRES CEA IM 2 and detect the expression of the plasmid in vitro. Methods:Using cloning technique,the cDNA fragments of CEA gene and mouse molecule IL 2 gene were constructed into the mammalian co expression plasmid vector pIRES.The inserted target genes in the mammalian co expression plasmid were verified by nucleotide sequencing.C 26 cell line was transfected with this mammalian co expression plasmid using lipofecarnin reagent. The expression of CEA and IL 2 molecules were detected by ELISA technique.Results:The mammalian co expression plasmid pIRES CEA IL 2 was obtained by cloning technique. The nucleotide sequences of CEA gene and molecule IL 2 gene in this mammalian co expression plasmid had high homology with CEA standard strain (99.8%) and mouse IL 2 (99.9%) respectively. After transfection with this mammalian co expression plasmid, the CEA and IL 2 molecules were expressed in C 26 cells. Conclusions:The constructed mammalian co expression plasmid pIRES CEA IL 2 can express CEA and IL 2 molecules in vitro at the same time.