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Reagent Information
Name Ad-mCMV-Flt3L 
Reagent Type Vector 
Vector
Classifications
Adenovirus
Keywords
 
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 transgene for human soluble fms-like tyrosine kinase 3 ligand (Flt3L) under the control of the murine cytomegalovirus (mCMV) promoter. The Ftl3L transgene was subcloned into the first generation adenoviral shuttle plasmid pAL120 containing the mCMV promoter to make the plasmid pAL120-hsFlt3L. The adenoviral vector genome plasmid pJM17 (Microbix) and pAL120-hsFlt3L were co-transfected in 293 cells (Microbix) and the adenoviral vector Ad-mCMV-Flt3L was rescued by homologous recombination. Ad-mCMV-Flt3L underwent three successive rounds of plaque purification and then amplified on 293 cells. Ad-mCMV-Flt3L 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-Flt3L.doc
Sequence  
Grade Research 
Depositor Cedars-Sinai Medical Center, Los Angeles, CA  
References 1. Dewey RA, Morrissey G, Cowsill CM, Stone D, Bolognani F, Dodd NJ, Southgate TD, Klatzmann D, Lassmann H, Castro MG, Löwenstein PR. Chronic brain inflammation and persistent herpes simplex virus 1 thymidine kinase expression in survivors of syngeneic glioma treated by adenovirus-mediated gene therapy: implications for clinical trials. Nature Medicine . 1999 Nov;5(11):1256-63. PMID: 10545991. 2. 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 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 4. Ali S, Curtin JF, Zirger JM, Xiong W, King GD, Barcia C, Liu C, Puntel M, Goverdhana S, Lowenstein PR, Castro MG. Inflammatory and anti-glioma effects of an adenovirus expressing human soluble Fms-like tyrosine kinase 3 ligand (hsFlt3L): treatment with hsFlt3L inhibits intracranial glioma progression. Molecular Therapy . 2004 Dec;10(6):1071-84. PMID: 15564139. 5. Ali S, King GD, Curtin JF, Candolfi M, Xiong W, Liu C, Puntel M, Cheng Q, Prieto J, Ribas A, Kupiec-Weglinski J, van Rooijen N, Lassmann H, Lowenstein PR, Castro MG. Combined immunostimulation and conditional cytotoxic gene therapy provide long-term survival in a large glioma model . Cancer Research . 2005 Aug 15;65(16):7194-204. PMID: 16103070. 6. Curtin JF, King GD, Barcia C, Liu C, Hubert FX, Guillonneau C, Josien R, Anegon I, Lowenstein PR, Castro MG. Fms-like tyrosine kinase 3 ligand recruits plasmacytoid dendritic cells to the brain . Journal of Immunology . 2006 Mar 15;176(6):3566-77. PMID: 16517725. 7. Larocque D, Sanderson NSR, Bergeron J, Curtin JF, Girton J, Wibowo M, Bondale N, Kroeger KM, Yang J, Lacayo LM, Reyes KC, Farrokhi C, Pechnick, Castro MG, Lowenstein PR. Exogenous fms-like tyrosine kinase 3 ligand overrides brain immune privilege and facilitates recognition of a neo-antigen without causing autoimmune neuropathology . The Proceedings of the National Academy of Sciences, USA . 2010 Aug 10;107(32):14443-8. 
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 descretion of the investigator's Institutional Biosafety Committee that may require a different level of containment. 
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