Title: Construction and expression of eukaryotic plasmid with dual-promoters and dual-reporter genes
Abstract: Objective To construct a eukaryotic plasmid with dual-promoters and dual-reporter genes,and express in 293T cells.Methods EGFP and DsRed genes were amplified by PCR using plasmids pEGFP-N1 and pDsRed-N1 as templates respectively and inserted into expression vector pCI.The constructed recombinant plasmids pCI-EGFP and pCI-DsRed were digested with BglⅡand BamHⅠthen spliced to construct recombinant eukaryotic expression vector pCI-cmvDsRed-cmvEGFP containing dual-promoters and dual-reporter genes.293T cells were transfected with plasmid pCI-cmvDsRed-cmvEGFP,in which the expressions of EGFP and DsRed were observed by fluorescent microscopy,Western blot and flow cytometry.Results Recombinant plasmid pCI-cmvDsRed-cmvEGFP was constructed correctly as proved by restriction analysis and sequencing,in which two promoters were linked in the same direction.Both EGFP and DsRed were expressed correctly at the same time.Conclusion A eukaryotic plasmid with dual-promoters and dual-reporter genes was successfully constructed and expressed in 293T cells.
Publication Year: 2012
Publication Date: 2012-01-01
Language: en
Type: article
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