Long-Term Stability Study and Topology Analysis of Plasmid DNA By Capillary Gel Electrophoresis
نویسندگان
چکیده
LEVEL: INTERMEDIATE T he topology of plasmid DNA can be monitored with capillary gel electrophoresis (CGE) (1, 2). An improved CGE technology was developed at PlasmidFactory to routinely determine the content of different plasmid forms that appear during cultivation and purification in DNA production runs. CGE is a useful tool for supporting process development in plasmid DNA and minicircle production, in drug delivery research, and in plasmid storage and stability evaluation studies. Long-term storage of the lacZ (beta-galactosidase)–expressing pCMVß plasmid over a period of 13 months revealed that stable storage conditions at –80 °C (rather than storage at 4 °C) prevent an increase in open circular (oc) plasmid topology deriving from the degraded, supercoiled, covalently closed circular (ccc) form. Physical data were supported by in vivo gene transfer data, using jet-injection into different tumor models grown subcutaneously in mice. Additionally, plasmid pCMV-S2S produced for clinical applications has been stored at –20° C and monitored for more than five years with no detectable decrease of quality (no topology changes).
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