©FUNPEC-RP www.funpecrp.com.br Genetics and Molecular Research 11 (1): 775-789 (2012) Production of human factor VIII-FL in 293T cells using the bicistronic MGMT(P140K)- retroviral vector A.M. Fontes 1,2 , F.U.F. Melo 1 , L.J. Greene 1 , V.M. Faça 1 , Y. Lin 3 , S.L. Gerson 3 and D.T. Covas 1,2 1 Hemocentro de Ribeirão Preto, Instituto Nacional de Ciência e Tecnologia em Células Tronco e Terapia Celular, Hospital das Clínicas, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil 2 Departamento de Clínica Médica and Departamento de Genética, Faculdade de Medicina de Ribeirão Preto, Universidade de São Paulo, Ribeirão Preto, SP, Brasil 3 Case Comprehensive Cancer Center, Case Western Reserve University and Seidman Cancer Center, University Hospitals Case Medical Center, Cleveland, OH, USA Corresponding author: A.M. Fontes E-mail: fontesam@hemocentro.fmrp.usp.br Genet. Mol. Res. 11 (1): 775-789 (2012) Received February 2, 2012 Accepted March 2, 2012 Published March 22, 2012 DOI http://dx.doi.org/10.4238/2012.March.22.8 ABSTRACT. Hemophilia A is the most common X-linked bleeding disorder; it is caused by deiciency of coagulation factor VIII (FVIII). Replacement therapy with rFVIII produced from human cell line is a major goal for treating hemophilia patients. We prepared a full-length recombinant FVIII (FVIII-FL), using the pMFG-P140K retroviral vector. The IRES DNA fragment was cloned upstream to the P140K gene, providing a 9.34-kb bicistronic vector. FVIII-FL cDNA was