Tracking and Quantitation of Retroviral-Mediated Transfer Using a Completely Humanized, Red-Shifted Green Fluorescent Protein Gene

R.R. Muldoon, J.P. Levy, S.R. Kain, P.A. Kitts, C.J. Link
1997 BioTechniques  
We have developed murine retroviral vectors (RVs) containing an optimized green fluorescent protein (GFP) gene to study retroviral gene transfer and expression in living cells. We used the codon "humanized", "red-shifted" GFP gene, hGFP-S65T , a gain of function variant of the wild-type GFP from the jellyfish Aequorea victoria . We cloned the hGFP-S65Tgene into the RV plasmid pLNCX (pLNChG65T). A stable amphotropic RV-producer cell line (VPC), designated LNChG65T VPC, was generated that
more » ... erated that exhibited bright fluorescence in greater than 95% of the cells. Human A375 melanoma cells and IGROV ovarian carcinoma cells transduced from LNCh -G65T VPC demonstrated high levels of fluorescence. The expression of a single integrated hGFP-S65Tgene in eukaryotic cells provides a powerful tool to study gene transfer, expression and functional studies in vitro and in vivo.
doi:10.2144/97221rr03 pmid:8994663 fatcat:frntzb2p2rf2xoj6hdjnl3cspq