TechnologyThe novel "RBPS-IVR Technology" is based on the coordinated consecutive steps of in vivo recombination, in vivo restriction and the subsequent chromatographic purification to remove residual impurities. The inducible sequence specific recombination process results in mc-DNA containing the gene of interest for the desired applications and miniplasmids carrying the unwanted bacterial backbone sequences. Triggered by this recombination based minicircle-DNA production step the expression of an endonuclease is initiated. The restriction sites of this endonuclease are located on the parental plasmid and - after recombination/minicircle production - on the miniplasmid. Upon expression of the endonuclease, plasmids carrying the corresponding sites are linearised and subsequently further degraded by host exonucleases. Therefore, after recombination and in vivo restriction/degradation the host cell harbours almost exclusively minicircle-DNA. Remaining trace amounts of residual parental plasmids can subsequently be removed by routine DNA chromatography techniques optimised for minicircle production.
Developmental statusThe RBPS-Technology has been successfully tested and scaled up using GMP compliant high cell density fermentation and chromatographic purification processes established in a common project with
BIASeparations, Eurogentec and the Ludwig Boltzmann Institute for Experimental and Clinical Traumatology.