Integration of two plasmid DNA (pDSRED, pEGFP) in one biopolymer multilayer system and its time-dependent expression in the cytoplasm

• Main Author: Oh, Bernice Hui Lin
• Other Authors: Bjoern Holger Neu
• Format: Final Year Project
• Language: English
• Published: 2009
• Subjects: DRNTU::Engineering::Chemical engineering::Biochemical engineering; DRNTU::Science::Biological sciences::Biophysics
• Online Access: http://hdl.handle.net/10356/16389
• Institution: Nanyang Technological University

In recent years there has been great interest in the study of polyelectrolyte multilayer microcapsules fabricated using the layer-by-layer technique. It offers great potential as a drug delivery system as the release profile and quantity of incorporated therapeutic agents can be controlled. In this study, two different types of plasmid DNA (pEGFP and pDsRed) were incorporated as layer components in a multilayered assembly fabricated by the layer by layer electrostatic adsorption of biodegradable biopolymers (protamine sulfate and dextran sulfate) onto negatively charged Silica core. The distance between the 2 plasmids was varied and their stability and properties were checked using flow cytometry and confocal microscopy. It was established that a minimum distance of three polyelectrolytes layers was required between two plasmid layers for stability. The expression rate of fluorescence proteins was monitored over a period of 3 days after incubating the particles with HEK293T cells. pDsRed incorporated in the outer layers were expressed earlier than pEGFP in the inner depth of the multilayer system. A higher expression rate of both proteins was observed with the system having a closer distance between the 2 incorporated pDNA. As the time dependent degradation profile is directly correlated to the rate of expression, it can be concluded that the defoliation of the multilayer proceeds progressively from the outer layers to the inner layers. The insights obtained from this project provide high potential in the therapeutic field for the further development of multiple-drug carriers with sustained release-kinetics.