肿瘤抗原P1A与细胞因子mGM-CSF共表达载体的构建和表达-东南大学学报医学版

肿瘤抗原P1A与细胞因子mGM-CSF共表达载体的构建和表达

单位：东南大学基础医学院病原生物学及免疫学系,江苏,南京,210009

分类号：R730.51, R392.1, Q786

出版年·卷·期（页码）：2004·23·第三期（184-188）

摘要：

目的:构建并鉴定细胞因子mGM-CSF和肿瘤特异抗原P1A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/P1A中扩增出P1A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pRSC-P1A重组质粒P1A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P1A基因和mGM-CSF基因的表达.结果:经酶切鉴定和DNA测序证实mGM-CSF和P1A重组质粒构建正确,并在转染此质粒的7721细胞中检测出了P1A和mGM-CSF基因的表达.结论:成功构建mGM-CSF和P1A的共表达载体,为研制新型肿瘤疫苗奠定了基础.

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.

参考文献：

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