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为构建带有荧光标签的汉滩病毒受体整合素β3与细胞因子TNFRⅠ的稳定转染细胞系,构建目的基因的真核表达载体pIRES2-EGFP-β3和pIRES2-DsRed-TNFRⅠ(EC),并转染CHO细胞,直接荧光观察报告基因和荧光定量RT-PCR检测目的基因mRNA表达。结果转染重组质粒pIRES2-EGFP-β3的细胞可检测到荧光报告基因和目的基因β3 mRNA的表达,转染pIRES2-DsRed-TNFRⅠ(EC)的细胞可检测到荧光报告基因和目的基因TNFRⅠmRNA的表达。结果表明成功地构建了真核表达整合素β3与TNFRⅠ的稳定细胞系。
To construct eukaryotic expression vectors pIRES2-EGFP-β3 and pIRES2-DsRed-TNFRⅠ (EC) of target gene and construct a stable transfected cell line with fluorescently tagged Hantaan virus receptor integrinβ3 and cytokine TNFRⅠand Transfection of CHO cells, direct fluorescent reporter gene and fluorescent quantitative RT-PCR detection target gene mRNA expression. Results The expression of fluorescent reporter gene and target gene β3 mRNA was detected in cells transfected with recombinant plasmid pIRES2-EGFP-β3. The expression of fluorescent reporter gene and target gene TNFRⅠmRNA was detected in cells transfected with pIRES2-DsRed-TNFRⅠ (EC) . The results showed that a stable cell line expressing integrin β3 and TNFR Ⅰ was successfully constructed.