肿瘤抗原P1A与细胞因子mGM-CSF共表达载体的构建和表达

目的：构建并鉴定细胞因子mGM-CSF和肿瘤特异抗原PIA共表达载体，为研究新型的肿瘤疫苗提供新的策略。方法：以PCR方法从pCI-neo／PIA中扩增出PIA编码基因片段，构建于pRSC质粒上；再从pORF-mGM-CSF中扩增出mGM-CSF基因片段．插入pRS-PIA重组质粒PIA基因的上游。以酶切和DNA测序进行鉴定。并将鉴定过的重组质粒以脂质体法转染7721细胞．用RT-PCR法鉴定转染细胞中PIA基因和mGM-CSF基因的表达。结果：经酶切鉴定和DNA测序证实mGM-CS17和PIA重组质粒构建正确，并在转染此质粒的7721细胞中检测出了PIA和mGM-CSF基因的表达。结论：成功构建mGM-CSF和PIA的共表达载体，为研制新型肿瘤疫苗奠定了基础。

Construction and expression of an coexpression vector of pRSC-mGM-CSF/P1A

Objective To construct the coexpressing vector of pRSC-mGM-CSF/P1A containing genes of P1A and mGM-CSF for the purpose of developing tumor vaccine.Methods P1A gene was amplified from pCI-neo/P1A vector by PCR and was ligated into vector pRSC to form pRSC-P1A recombinant.The mGM-CSF gene which was obtained from pORF-mGM-CSF vector by PCR was inserted to the upstream of the P1A in pRSC-P1A recombinant to develop a pRSC-mGM-CSF/P1A tumor vaccine constructs.After the identified recombinant plasmids were transformed into cell 7721 with lipofectamine,the expression of P1A and mGM-CSF in the cells were determined with RT-PCR. Results The pRSC-mGM-CSF/P1A tumor vaccine constructs was identified by restrictive digestion and sequencing methods respectively, and the expression of P1A gene and mGM-CSF gene could be detected in the transformed 7721 cells.Conclusion The recombinant pRSC-mGM-CSF/P1A has been successfully constructed,and it provides a good foundation for developing novel tumor vaccine.