Brief Reports
Copyright ©2005 Baishideng Publishing Group Inc. All rights reserved.
World J Gastroenterol. Apr 28, 2005; 11(16): 2497-2501
Published online Apr 28, 2005. doi: 10.3748/wjg.v11.i16.2497
Effective siRNA targets screening for human telomerase reverse transcriptase
Yun Xia, Ru-Xian Lin, Su-Jun Zheng, Ying Yang, Xiao-Chen Bo, Dao-Yin Zhu, Sheng-Qi Wang
Yun Xia, Su-Jun Zheng, Ru-Xian Lin, Ying Yang, Xiao-Chen Bo, Sheng-Qi Wang, Beijing Institute of Radiation Medicine, Beijing 100850, China
Yun Xia, Dao-Yin Zhu, Department of Microbiology and Immunology, Chongqing Medical University, Chongqing 400016, China
Author contributions: All authors contributed equally to the work.
Supported by the National Natural Science Foundation of China, No. 30371662
Correspondence to: Professor Sheng-Qi Wang, Beijing Institute of Radiation Medicine, No. 27 Taiping Road, Beijing 100850, China. sqwang@nic.bmi.ac.cn
Telephone: +86-10-66932211 Fax: +86-10-66932211
Received: March 20, 2004
Revised: March 21, 2004
Accepted: April 29, 2004
Published online: April 28, 2005
Abstract

AIM: To study the inhibitory effects of siRNAs targeting different hTERT sequences and to screen the effective siRNA sequence.

METHODS: Five double-stranded siRNAs targeting coding and non-coding regions of hTERT gene were designed and synthesized by T7 transcription system in vitro. siRNA4 sequence was screened by full length gene targeting technique and the rest of the siRNA sequences were selected randomly. After being purified by ethanol precipitation, the siRNAs were transfected to the human hepatocellular carcinoma cell (HepG2) by LipofectamineTM2000. At 48-72 h after siRNAs transfection, MTT assay, RT-PCR and Western-blot were applied to evaluate the effects of siRNAs on cell growth, mRNA and protein expression level of hTERT gene, respectively.

RESULTS: Compared to the control cells, the cells treated with the five double-stranded siRNAs exhibited different degrees of inhibition of cell proliferation in a dose-dependent manner. siRNA2 and siRNA4, exhibited obvious effects of inhibiting hTERT mRNA and protein expression in HepG2 cells.

CONCLUSION: siRNAs targeting different hTERT sequences have significantly various inhibitory effects on hTERT gene expression. The siRNA sequence screened by full length gene targeting technique has comparable inhibitory effect with the rest siRNA sequences screened by random selection, suggesting that siRNAs and antisense oligonucleic acids may have the same effective target sites. Compared with chemical synthesis method, synthesizing double-stranded siRNA by T7 transcription system in vitro is a rapid, simple, and inexpensive method suitable for screening high-effect siRNA targeting site for specific gene.

Keywords: siRNA targets; hTERT