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Reagent Information
Name Ad-mCMV-Teton 
Reagent Type Vector 
Cell Line Origin  
Short Description  
Long Description This replication-defective, first generation adenoviral vector is based on human adenovirus serotype 5, with deletions in its E1a and E3 viral encoding regions. This vector encodes the transgenes for mutant tetracycline-dependent transactivator (rtTA2SM 2 ) and transcriptional silencer carrying plasmid tTSKid under the control of the murine cytomegalovirus (mCMV) promoter. The rtTA2SM 2 and tTSKid are separated by an IRES sequence. The rtTA2SM 2 -IRES-tTSKid expression cassette was excised from the plasmid pMTIKA (Addgene plasmid 22061) and subcloned into the first generation adenoviral shuttle plasmid E1sp1a to make the plasmid E1sp1a-rtTA2SM 2-IRES-tTSKid. The adenoviral vector genome plasmid pJM17 (Microbix) and E1sp1a-rtTA2SM 2-IRES-tTSKid were co-transfected in 293 cells (Microbix) and the adenoviral vector Ad-mCMV-TetOn was rescued by homologous recombination. Ad-mCMV-TetOn underwent three successive rounds of plaque purification and then amplified on 293 cells. Ad-mCMV-TetOn was purified from the 293 cell lysate by cesium chloride purification followed by three rounds of dialysis in 10mM Tris, ph 8.0.  
Vector Map Ad-mCMV-TetOn.doc
Grade Research 
Depositor Cedars-Sinai Medical Center, Los Angeles, CA  
References 1. Gerdes CA, Castro MG, Löwenstein PR. Strong promoters are the key to highly efficient, noninflammatory and noncytotoxic adenoviral-mediated transgene delivery into the brain in vivo . Mol Ther. 2000 Oct;2(4):330-8. PMID: 11020348 3. Appleby CE, Kingston PA, David A, Gerdes CA, Umaña P, Castro MG, Lowenstein PR, Heagerty AM. A novel combination of promoter and enhancers increases transgene expression in vascular smooth muscle cells in vitro and coronary arteries in vivo after adenovirus-mediated gene transfer . Gene Ther . 2003 Sep;10(18):1616-22. PMID: 12907954 3. Xiong W, Goverdhana S, Sciascia SA, Candolfi M, Zirger JM, Barcia C, Curtin JF, King GD, Jaita G, Liu C, Kroeger KM, Ng P, Palmer D, Agadjanian H, Medina-Kauwe L, Lowenstein PR, Castro MG. Regulatable Gutless Adenovirus Vectors are Capable of Regulating Transgene Expression in the Brain in the Presence of an Immune Response Against Adenoviruses . Journal of Virology 2006 Jan;80(1):27-37. PMID: 16352528 . 4. Candolfi M, Curtin JF, Xiong W, Kroeger KM, Liu C, Rentsendorj A, Agadjanian H, Medina-Kauwe L, Palmer D, Ng P, Lowenstein PR, Castro MG. Effective High Capacity Gutless Adenoviral Vectors Mediated Transgene Expression in Human Glioma Cells . Molecular Therapy 2006 Sep;14(3):371-381. PMID: 16798098 5. Xiong W, Candolfi M, Kroeger KM, Puntel M, Mondkar S, Larocque D, Liu C, Curtin JF, Palmer D, Ng P, Lowenstein PR, Castro MG. Immunization against the transgene but not the TetON switch reduces expression from gutless adenoviral vectors in the brain . Molecular Therapy 2008 Feb;16(2):343-51. PMID: 18180781 2. Candolfi M, Xiong W, Yagiz K, Liu C, Muhammad AK, Puntel M, Foulad D, Zadmehr A, Ahlzadeh GE, Kroeger KM, Tesarfreund M, Lee S, Debinski W, Sareen D, Svendsen CN, Rodriguez R, Lowenstein PR, Castro MG. Gene therapy-mediated delivery of targeted cytotoxins for glioma therapeutics . The Proceedings of the National Academy of Sciences USA . 2010 Oct 28. PMID: 21030678  
Biosafety The Indiana University Vector Production Facility works with this line under BL-2 containment but the level of containment required of any cell line or vector is at the discretion of the investigator's Institutional Biosafety Committee that may require a different level of containment