Probing DNA assembly into nanoparticles with short DNA
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Probing DNA assembly into nanoparticles with short DNA Preethi L. Chandran1,2, Emilios K. Dimitriadis1, and Ferenc Horkay1 1 Section on Tissue Biophysics and Biomimetics, NICHD, 2 Laboratory of Bioengineering and Physical Science, NIBIB, National Institutes of Health, Bethesda, MD 20892, U.S.A. ABSTRACT DNA is an anionic polyelectrolyte, which occupies a large volume in salt free solution due to the coulomb repulsion between the charged groups. In the presence of high valence cations, DNA condenses into nanoparticles. DNA nanoparticles have generated a lot of interest as a preferred vehicle for delivering therapeutic DNA in gene therapy. The efficiency of gene delivery is determined by stability and compactness of the particles. However not much is known about the organization of DNA within the particles. The large polymer cations condense DNA rapidly, with no distinct intermediate stages that give insight into the arrangement of DNA within the nanoparticle. In our work, we form nanoparticles with short DNA strands to slow down the condensation process. The polymer cation is polyethyleneimine with grafted sugar moieties. Distinct intermediate stages are observed with Atomic Force Microscopy. The assembly occurs via the formation of fiber condensates, which appear to be the unit of DNA condensation. Nanoparticles form by compaction of interweaving networks of fiber condensates. INTRODUCTION DNA is a highly negatively charged polymer. It is semirigid (persistence length of 50 nm) and self-repelling, occupying an expanded volume. However in the presence of polycations (3 positive charges or higher), DNA can be condensed to less than 1000 times its free volume; allowing several micrometers of DNA be packed into viruses and bacteria of nanometer size. DNA condensation with polycations has generated a lot of interest both from a basic sciences viewpoint to understand DNA packaging in viruses, but also for packaging DNA into nanoparticles for delivery into cells. The human genome project and the prospect of combating diseases by delivery of therapeutic genes has fuelled a lot of research into using DNA nanoparticles as gene delivery agents [1]. Commonly used cationic polymers like polyethyleneimine condense DNA into stable, compact particles [1, 2]. The net positive charge on the nanoparticles allows attractive interaction with the negatively charged cell surface [2]. The nanoparticle size is on the order of virus particles, and is taken up within the cell by the preferred endocytosis machinery. The buffering action of polyethyleneimine protects the DNA from the acidic environment of the endocytosis vesicles. It is increasingly appreciated that small changes in the DNA organization within the nanoparticles have large influences on the biological activity of the particles [3]. Therefore a systematic study of DNA organization within nanoparticles and its correlation with biological activity is warranted. However, for DNA lengths typically studied (> 10 kb) and for condensation with polymer cations, nanoparticles form rapidly
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