title: Doxycycline-dependent inducible and reversible RNA interference mediated by a single lentivirus vector.
abstract: RNA interference has been applied to the development of a method of silencing genes of interest. Short hairpin RNA (shRNA) expression vectors are useful in driving gene-silencing. Standard shRNA vectors produce a knockdown phenotype soon after transduction. An alternative strategy for conditional gene knockdown would be useful to investigate gene functions in a time-dependent manner. In this study, we developed an inducible gene knockdown system based on lentivirus-mediated gene transfer. A single lentivirus vector capable of inducible expression of a designed microRNA-based shRNA was generated using a tetracycline-dependent transactivation system. The lentiviral vector facilitated doxycycline-dependent inducible knockdown specific to the target gene. Withdrawal of doxycycline after transient treatment resulted in complete recovery of target gene expressions to normal levels. Thus the single lentiviral vector developed in this study should be a powerful tool for doxycycline-dependent inducible and reversible RNA interference in molecular genetic studies.
authors.affiliation: Functional Genomics Core Laboratory, Ehime University Proteo-Medicine Research Center (ProMRes), Ehime, Japan. natsukim@m.ehime-u.ac.jp
authors.forename: Natsuki
authors.initials: N
authors.lastname: Matsushita
authors.forename: Sachi
authors.initials: S
authors.lastname: Matsushita
authors.forename: Satoshi
authors.initials: S
authors.lastname: Hirakawa
authors.forename: Shigeki
authors.initials: S
authors.lastname: Higashiyama
mesh_terms: D004318:Doxycycline / Q000494:pharmacology*; D055785:Gene Knockdown Techniques / Q000379:methods*; D018773:Genes, erbB-1 / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics*; D057809:HEK293 Cells; D006801:Humans; D016086:Lentivirus / Q000235:genetics*; D061986:MCF-7 Cells; D034622:RNA Interference / Q000187:drug effects*; D015533:Transcriptional Activation / Q000187:drug effects; D019076:Transgenes / Q000235:genetics
chemical_list: D004318:Doxycycline