RNA-Seq of H1299 cells transfected with different types of bi-directional plasmid to better understand the transcriptional changes occurring with the different design frameworks

Our previous work has shown that exogenous DNA impose a burden to the host cells due to the finite amount of transcriptional and translational resources, resulting in a limiting productivity capacity. We thus designed context-aware genetic networks in which miRNAs were successfully exploited as burden mitigators, and observed upregulation of protein expression in the targeted cells. We co-transfected H1299, the testbed of our previous studies with a bi-directional plasmid encoding EGFP and mKate, either in an open loop circuit architecture (OL) that lack miRNA regulation, or in the form of a miRNA-iFFL, placing miR31 target sites, highly expressed in this cell line, in the 3' (iFFL-3') or 5'(iFFL-5') UTR of mKate (Fig. 2a). We next sorted the cells that were either non-transfected (NTr) or transfected (Tr) according to fluorescence expression, to compare the OL that are more sensitive to burden, with iFFL-3' and iFFL-5' architectures.