关键词： 5-FC 5-fluorocytosine CD Ct DMEM Dulbecco's modified Eagle's Medium GCV Gastric cancer Gene therapy MAR MTT Matrix attachment region PCR PI Suicide gene Survivin promoter TK cytosine deaminase ganciclovir matrix attachment region methyl thiazolyl tetrazolium polymerase chain reaction propidium iodide qPCR quantitative real-time PCR threshold cycle number thymidine kinase

Mesh ： Apoptosis / genetics Cell Line, Tumor Cell Survival / genetics Genes, Transgenic, Suicide Genetic Therapy / methods Genetic Vectors / genetics Humans Inhibitor of Apoptosis Proteins / genetics Matrix Attachment Regions / genetics Plasmids / genetics Promoter Regions, Genetic Stomach Neoplasms / genetics pathology therapy Survival Rate Survivin

来  源：   DOI：10.1016/j.gene.2013.10.064   PDF(Sci-hub)

Abstract：
This work aimed to study a novel transgenic expression system of the CD/TK double suicide genes enhanced by the nuclear matrix attachment region (MAR) for gene therapy. The recombinant vector pMS-CD/TK containing the MAR-survivin promoter-CD/TK cassette was developed and transfected into human gastric cancer SGC-7901 cells. Expression of the CD/TK genes was detected by quantitative real-time PCR (qPCR) and Western blot. Cell viability and apoptosis were measured using the methyl thiazolyl tetrazolium (MTT) assay and flow cytometry. When the MAR fragment was inserted into the upstream of the survivin promoter, the qPCR result showed that the expression of the CD/TK genes significantly increased 7.7-fold in the transgenic SGC-7901 cells with plasmid pMS-CD/TK compared with that without MAR. MTT and flow cytometry analyses indicated that treatment with the prodrugs (5-FC+GCV) significantly decreased the cellular survival rate and enhanced the cellular apoptosis in the SGC-7901 cells. The expression of the CD/TK double suicide genes driven by the survivin promoter can be enhanced by the MAR fragment in human gastric cancer cells.
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