Abstract:

BACKGROUND: Human telomerase reverse transcriptase (hTERT) closely correlates with c-myc in laryngeal squamous cell carcinoma.
OBJECTIVE: To construct recombinant adenovirus vector of hTERT containing c-myc promoter and to observe the effects of expressed hTERT on cell growth.
METHODS: Using the method of reverse transcription in artificial synthesis to design the gene synthesis primer and target fragment ashTERT (322 bp) was obtained using overlapping PCR. The target fragment ashTERT was cloned into the vector pUC57(2.7 kb) and then heat-shock transformed into the competent cells E.coli. With Bacteria inspection, PCR, identification of positive clones, plasmid pUC57-ashTERT was extracted and sequenced. The target gene fragments were subcloned into the shuttle vector pshuttle-CMVneo and then the recombinant adenovirus plasmid pAd-ashTERT was constructed. Subsequently, the recombinant adenovirus plasmid pAd-ashTERT was transformed into the 293A cells for package, identification and titer determination of adenovirus.
RESULTS AND CONCLUSION: pUC57-ashTERT plasmid was extracted and its sequence corresponded to sequencing requirement. Recombinant adenovirus vector of hTERT with relatively strong ability of infection was successfully constructed after pUC57-ashTERT plasmid was transformed into 293 cells.