癌胚抗原与γ-干扰素真核双表达质粒的构建及体外表达

Construction of Mammalian Co-expression Plasmid pIRES-CEA-IFN γ and Its Expression in vitro

目的 :构建癌胚抗原 (CEA)与γ 干扰素 (γ IFN)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γ IFN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γ IFN的表达。结果 :核酸序列测定证实本实验所构建的质粒正确 ,该双表达质粒在体外转染细胞后可表达CEA和γ IFN分子。结论 :实验所构建的pIRES CEA γIFN双表达质粒能在体外同时表达CEA和γ

Objective To construct the mammalian co-expression plasmid pIRES-CEA-IFN γ and detect the expression of the plasmid in vitro.Metheds Using cloning technique,the cDNA fragments of CEA gene and mouse molecule γ-IFN 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 γ-IFN molecules were detected by ELISA technique.Results The mammalian expression plasmid pIRES-CEA-IFN γ was obtained by cloning technique.The nucleotide sequences of CEA gene and molecule IFN γ gene in this mammalian co-expression plasmid had high homology with CEA standard strain(99.8%)andmouseγ IFN (99 99% )respectively .Aftertransfectionwiththismammalianco expressionplasmid ,the CEAandγ IFNmoleculeswereexpressedinC 2 6cells .Conclusion Theconstructedmammalianco expres sionplasmidpIRES CEA IFNγcanexpressCEAandγ IFNmoleculesinvitroatthesametime .