Liposomes for DNA Nanotechnology: Preparation, Properties, and Applications - DNA Nanotechnology: From Structure to Function - page 64

Fig. 4.4 Reactive lipid head groups in N-PDP-PE ( a )andMPB-PE( b ) for conjugating with DNA

4.5.5 Removal of Free DNA

As described above, certain lipid-modified DNA can quantitatively insert itself
into lipid bilayer, omitting the need for purification. For other cases, a separation
step needs to be performed to remove free DNA. A popular method to achieve
this is to use short gel permeation columns, where DNA-liposome conjugates
migrate faster than free DNA. Commonly used short columns are unlikely to have
very high separation efficiency though. In addition, the liposome samples might
be significantly diluted if longer columns are used. Interestingly, at high DNA
density, many DNA sequences can induce self-aggregation of liposomes [ 36 ]. This
is because of associated DNA being partially complementary to itself combined with
multiple copies of DNA displayed on the liposomes [ 38 ]. This aggregation is further
facilitated by storage at low temperature (e.g., 4 ı C) in the presence of high salt
(e.g., 500 mM NaCl), which favors DNA hybridization. The aggregated liposomes
can be easily harvested by a brief centrifugation at 4 ı C. For some DNA sequences,
self-aggregation does not occur readily. In these cases, ultracentrifugation can be
performed at 100,000 rpm. Regardless of the method employed, removal of free
DNA is critical for many analytical and biomedical applications.