Atomic force microscopy reveals the assembly of potential DNA "nanocarriers" by poly-L-ornithine.
Atomic force microscopy (AFM) has been used to visualize the process of condensation of plasmid DNA by poly-L-ornithine on mica surface. AFM images reveal that the transition of negatively charged DNA to condensed nanoparticles on addition of increasing amounts of positively charged poly-L-ornithine (charge ratio (Z+/Z-) varied between 0.1 and 1) at a wide range of DNA concentrations (3-20 ng/microl) occurs through formation of several distinct morphologies. The nature of the complexes is strongly dependent on both the charge ratio and the DNA concentration. Initiation of condensation when the concentration of DNA is low (approximately 3-7 ng/microl) occurs possibly through formation of monomolecular complexes which are thick rod-like in shape. On the contrary, when condensation is carried out at DNA concentrations of 13-20 ng/microl, multimolecular structures are also formed even at low charge ratios. This difference in pathway seems to result in differences in the extent of condensation as well as size and aggregation of the nanoparticles formed at the high charge ratios. To the best of our knowledge, this is the first direct single molecule elucidation of the mechanism of DNA condensation by poly-L-ornithine. Cationic poly-aminoacids like poly-L-ornithine are known to be efficient in delivery of plasmid DNA containing therapeutic genes in a variety of mammalian cell lines by forming condensed "nanocarriers" with DNA. Single molecule insight into the mechanism by which such nanocarriers are packaged during the condensation process could be helpful in predicting efficacy of intracellular delivery and release of DNA from them and also provide important inputs for design of new gene delivery vectors.
Mann
Anita
A
Institute of Genomics and Integrative Biology, Mall Road (near Jubilee Hall), Delhi 110 007, India.
Khan
Meraj Alam
MA
Shukla
Vasundhara
V
Ganguli
Munia
M
D000538:Aluminum Silicates / Q000737:chemistry; D004247:DNA / Q000737:chemistry*; D024202:Electrophoretic Mobility Shift Assay; D018625:Microscopy, Atomic Force; D053769:Nanocapsules / Q000737:chemistry*; D009690:Nucleic Acid Conformation; D010455:Peptides / Q000737:chemistry*; D010957:Plasmids / Q000737:chemistry
D000538:Aluminum Silicates; D053769:Nanocapsules; D010455:Peptides; C008973:polyornithine; D004247:DNA; C011934:mica