Periodic Reporting for period 1 - APPLIC-ION (A universally applicable system for direct readout of eukaryotic cell transgenesis with a novel integration-coupled gene expression switch)
Periodo di rendicontazione: 2021-04-01 al 2022-09-30
Many biological, biotechnological and biomedical applications rely on stable integration of exogenous transgenes in the genome of transfected eukaryotic host cells. Due to their low cost, easy production and simplicity of use, DNA plasmids are increasingly applied for this purpose as an alternative to viral particles. However, stable transgenesis with these vectors remains slow and complex due to the necessity to select stable transfected cells, a weeks-long procedure with risks of toxicity or epigenetic drift.
To solve these issues, we have recently introduced a novel type of genetic switch that conditions DNA transgene expression to incorporation into the host genome. This switch termed “iOn” opens the way to radically simpler transgenesis procedures through direct and accurate identification of integration events following cell transfection. In the Applic-iOn ERC PoC project, we have further developed this system to facilitate its use and apply it to new purposes. Among others, we have created and validated new single-component iOn vectors that facilitate the leakproof integration-dependent expression of virtually any genes of interest. We have also established iOn-based strategies to interfere with the expression of specific endogenous genes. These advances widen the range of applications of the iOn toolkit to most of those permitted by complex transgenic approaches and genetic systems, thus significantly increasing the technological readiness of this new system.
To solve these issues, we have recently introduced a novel type of genetic switch that conditions DNA transgene expression to incorporation into the host genome. This switch termed “iOn” opens the way to radically simpler transgenesis procedures through direct and accurate identification of integration events following cell transfection. In the Applic-iOn ERC PoC project, we have further developed this system to facilitate its use and apply it to new purposes. Among others, we have created and validated new single-component iOn vectors that facilitate the leakproof integration-dependent expression of virtually any genes of interest. We have also established iOn-based strategies to interfere with the expression of specific endogenous genes. These advances widen the range of applications of the iOn toolkit to most of those permitted by complex transgenic approaches and genetic systems, thus significantly increasing the technological readiness of this new system.