关键词: PTEN基因, 质粒, SCC-4细胞系, 基因转染

Abstract: Abstract:Objective To construct the eukaryotic expressing vector of PTEN gene and to express the gene in SCC-4 cell line and to provide theoretical foundation for gene therapy of tongue squamous cell carcinoma.Methods  Human total length of PTEN gene was obtained from HeLa cell line by RT-PCR and was cloned into pMD18-T to analyze the sequence.The correct PTEN was inserted into pEGFP-N1 vector.pEGFP-PTEN was identified by digestion with EcoRⅠ and XhoⅠ.Scc-4 cell line was transfeted by pEGFP-PTEN and the expression of PTEN was observed by fluorescence microscope and Western blotting.Results  A 1 200 bp fragment had been cloned into pEGFP-N1 vector which was further identified by sequencing and NCBI BLAST analysis.pEGFP-PTEN was expressed successfully in SCC-4 cell line.Western blotting showed the expression of PTEN protein in pEGFP-PTEN group was 1.07±0.15,there were significant differences compared with pEGFP-N1 group(0.62±0.11) and blank control group(0.57±0.08) (P<0.05).Conclusion The eukaryotic recombinant vector pEGFP-PTEN is constructed successfully and could be overexpressed in SCC-4 cell line.

Key words: PTEN gene, plasmid construction, SCC-4 cell line, gene transfection