Construction and expression of hTERT in eucaryotic expression vector

[Objective] To construct and express a human telomerase reverse transcriptase eukaryotic expression vector pEGFP-C3. [Methods] The genomic fragment of hTERT was amplified by RT-PCR and inserted into the pGEM-T Easy vector, the recombinant plasmid pGEM-T- hTERT and pEGFP-C3 were cleaved by restriction endonuclease HindⅢ and BamHI, subcloned hTERT into eukaryotic expression vector pEGFP-C3, The recombinant pEGFP-C3- hTERT was transfected into NIH3T3 by Lipofectamine method. The stable expressing NIH3T3-hTERT was established through G418 screening, and was observed by fluorescent converted microscope, and the expression level of hTERT mRNA was evaluated. [Results] Compared with hTERT gene in GenBank, the homology of the recombined expression vector pEGFP-C3-hTERT was 100% in nucleotide acid sequence. Green fluorescence was seen and high level expression of human TERT mRNA was checked in transfected NIH3T3. [Conclusion] pEGFP-C3-hTERT is constructed successfully. It provides the data for further study based on hTERT as target in anti-tumor gene immunity therapy.