Development of a tightly regulated U6 promoter for shRNA expression.

Lin X, Yang J, Chen J, Gunasekera A, Fesik SW, Shen Y
FEBS Lett. 2004 577 (3): 376-80

PMID: 15556613 · DOI:10.1016/j.febslet.2004.10.033

Short hairpin RNAs (shRNAs) have been used to achieve stable target knockdown in a variety of biological systems. Here, we report the development of a tightly regulated tetracycline-responsive human U6 promoter for shRNA expression. By engineering two copies of the tet operators flanking the TATA box of the human U6 promoter, we created a U6 promoter derivative (2O2) that exhibited much lower basal transcriptional activity compared with recently reported inducible pol III dependent promoters. As a consequence of its tighter regulation, the 2O2 system greatly improved the success rate in making inducible knockdown cell lines.

MeSH Terms (16)

Blotting, Western Cell Line, Tumor Cloning, Molecular Gene Expression Regulation Genes, Reporter Genetic Variation HeLa Cells Humans Luciferases Molecular Sequence Data Promoter Regions, Genetic RNA, Small Interfering Sequence Homology, Nucleic Acid TATA Box Tetracycline Transcription, Genetic

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