A eukaryotic replicative pUC-free minicircle expression vector is provided. The eukaryotic replicative pUC-free minicircle expression vector includes a pUC-free eukaryotic region sequence encoding a transgene of interest and comprising 5′ and 3′ ends and ii) a pUC-free spacer region of less than 500 basepairs in length linking the 5′ and 3′ ends of the eukaryotic region sequences and comprising a bacterial R6K replication origin having at least 95% sequence identity to SEQ ID NO: 11 or SEQ ID NO: 12 and a RNA selectable marker, the RNA selectable marker being an RNA-IN regulating RNA-OUT functional variant having at least 95% sequence identity to SEQ ID NO:20 or SEQ ID NO:22.
