JOURNAL OF SHANDONG UNIVERSITY (OTOLARYNGOLOGY AND OPHTHALMOLOGY)

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Construction of RNAi expressing plasmid vector of pSIRENshuttle for EGFR gene silencing

LUAN Jiangang1, LIANG Chuanyu2,WEN Yanjun3, LI Jiong3   

  • Received:2005-12-12 Revised:2005-12-20 Online:2006-02-25 Published:2006-02-25

Abstract: [ABSTRACT]Objective: To construct an expressing vector (pSIRENShuttleEGFR ) of RNAi in order to suppress gene expression of epidermal growth factor receptor (EGFR). Methods: Three 69 basepair oligos coding hairpin RNA which specifically aimed at EGFR gene were chemically synthesized and annealed, then inserted into the downstream of U6 promoter of vector to construct RNAi plasmid (pSIRENShuttleEGFRs). Sequence analysis was used to identify the correction of recombinant vector. RT-PCR was performed on the CNE cell line transfected by pSIRENShuttleEGFRs. Results: Sequencing analysis demonstrated that 69 bp had been inserted into expected site and the insertion sequence was exactly correct in recombinant pSIRENShuttleEGFRs vectors. It was showed that usingthe plasmid vector pSIRENEGFRs to induce RNAi suppressed the expression of endogenous EGFR in CNE human nasopharyngeal epidermal carcinoma cells. As a consequence, the mRNA of EGFR was decreased. Conclusion: RNAimediated inhibition of EGFR gene induced by shRNA expressing plasmid vector may constitute a useful approach in the treatment of human nasopharyngeal epidermal cancer.

Key words: epidermal growth factor, RNA, Receptor, messenger, RNA, interference, RNA

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