Biomod/2013/BU/results
Boston University
BIOMOD 2013 Design Competition
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Results
Two faces of our structure contain the helix ends that we hypothesized can be used as sites of functionalization. By extending the oligonucleotides that touch the ends we successfully created sites of functionalization that can be used to tag our structure with bioactive peptides. We extended each helix in the 5’ prime direction by 14 thymine residues. Peptide hybrids possessing the complementary 14 adenosine residues should hybridize with this structure to produce a polyvalent peptide display that will dictate the particle’s bioactivity. Our product successfully formed in high folding yield when the helix ends were extended from one face of the structure and from two faces of the structure. A screen of magnesium concentrations was performed to discern the optimal folding concentration. Agarose gel electrophoresis and TEM conclusively demonstrated that the nanostructure is folding properly. After optimization of buffer concentration and spin speeds, we were able to remove excess oligonucleotides using centrifugal dialysis filters. An example of the structures after purification is shown in Figure 1 below. Progress is ongoing towards synthesizing peptide oligonucleotide conjugates which will be used to functionalize the surface of the DNA nanoparticle. Cell penetrating peptides and blood-brain-barrier penetrating peptides will be used. Final products will be tested on cell cultures to determine the mechanism and extent of cell entry, and in live mice, to determine brain localization.
Add the attached picture with this caption:
Figure 1. This figure shows a TEM image of a sample of 20nM 2-sided 14T-extended DNA boxes spun six times at 14,000g in the 30kDa filter. As is evident, the boxes formed successfully at a high yield. Note: the direct magnification is 68,000x while the print magnification is 4010x at 7 mm.
