Pmxs ires gfp

Four liver cancer cell lines with different mutant states of p53 (HepG2, wild-type; Hep3B, null; Huh7, mutant; and PLC, mutant) were cultured in Dulbecco’s modified Eagle medium (DMEM) including 10% heat inactivated fetal bovine serum (FBS) at 37°C under a 5% humidified CO₂ atmosphere. Newborn human foreskin fibroblasts (human dermal fibroblasts, HDFs) and gp-293 cells were purchased from Globalstem Inc. (Gaithersburg, MD, USA) and Takara Bio USA Inc. (Mountain View, CA, USA), respectively, and maintained in DMEM containing 10% FBS. Four liver cancer cell lines used for reprogramming and HDFs at passage 11 were also used as normal cell controls of reprogramming.19 (link) The reprogrammed cells were grown on a mouse embryonic fibroblast feeder layer with human embryonic stem cells growth medium containing DMEM/F12, 20% knockout serum, 2 mM GlutaMAX, 0.1 mM nonessential amino acids, 0.1 mM β-mercaptoethanol, 50 U/mL penicillin, and 50 g/mL streptomycin. The plasmids used in this study were purchased from Addgene (pMXs-hklf4, pMXs-hoct4, pMXs-hsox2, pMXs-hmyc, and pCMV-VSV-G; Cambridge, MA, USA) and Cell Biolabs, Inc. (pMXs-IRES-GFP; Huntsville, AL, USA).

pMXs-IRES-GFP was purchased from Cell Biolabs (San Diego, CA, USA). pMSCV-IRES-GFP was a kind gift from Dario Vignali (Addgene plasmid 52107). pBApo-EF1a Pur was purchased from Takara Bio (Shiga, Japan). CSII-EF-MCS, CSII-EF-MCS-IRES2-Venus, pCAG-HIVgp, and pCMV-VSV-G-RSV-Rev, kind gifts from Dr. H. Miyoshi, were provided by the RIKEN BRC through the National BioResource Project of the MEXT/AMED, Japan (RDB04378, RDB04384, RDB04394, and RDB04393, respectively).