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[Construction of antisense telomerase hTERT and its effect on K562 cells]
Xiu-xiang Meng1, Ben-li Su, Li Jia
1Dalian Medical University, Dalian 116027, China.
Objectives:
To investigate whether antisense human telomerase reverse transcriptase (hTERT) could inhibit the activity of telomerase and the proliferation of K562 cells.
Methods:
The antisense plasmid was constructed by reverse insertion of hTERT PCR product into plasmid pLNCX-neo. Then the constructed plasmid was introduced into K562 cells by liposomes-mediated DNA transfection. The inhibition effects of telomerase on the proliferation of K562 cells were analyzed by MTT and colony formation assay, the telomerase activity of K562 cells by TRAP-PCR ELISA methods.
Results:
The growth rate of antisense hTERT transfected K562 cells was significantly lower than those of the controls, and the colony formation capacity of the transfected cells decreased significantly (P < 0.01), the colony number is (100.33 +/- 7.57)/10(3) cells, (92.67 +/- 5.86)/10(3) cells and (50.33 +/- 6.11)/10(3) cells for control K562 cells, K562 neo cells and antisense hTERT transfected HL60 cells, respectively. The telomerase activity of antisense hTERT transfected K562 cells was significantly inhibited.
Conclusion:
The expression of an antisense sequence to the mRNA sequence of telomerase protein subunit can inhibit the activity of telomerase, slow the cell growth and inhibit the capacity of colony formation of K562 cells.