A master thesis from Aalborg University
Compaction of DNA into an Origami-like Structure for Efficient Gene Delivery
Author(s)
Term
4. term
Education
Publication year
2022
Submitted on
2022-06-02
Pages
75 pages
Abstract
During this Master's thesis, it was attempted to design and produce a DNA origami-like structure, able to be in vivo transfected into mammalian cells and express a fluorescent protein. Initially, the pHAGE-EF1αL-eGFP-W plasmid was utilized, but its poor sequencing proved it unsuitable. Next, a DNA origami design, which had as a template the TagRFP gene of the pTagRFP-C vector, was created through the aid of caDNAno2 and Maya's vHelix plugin. The structure's stability was in silico tested with oxDNA2. Asymmetric PCR was used to create the scaffold DNA, while an attempt to produce ten ssDNA fragments, essential for the origami structure, was performed. The annealing of the structure was evaluated through AGE and AFM imaging. Finally, different DNA origami annealing protocols were tested for their transfection efficiency on HeLa cell cultures, along with a carrier-free pTagRFP-C vector and the vector mixed with poly-L-lysine.
Keywords
DNA Origami ; pTagRFP-C ; Gene Delivery ; Transfection ; HeLa Cells ; aPCR ; Asymmetric PCR ; PCR ; AGE ; Poly-L-Lysine ; Cholesterol ; oxDNA ; caDNAno ; Maya ; vHelix ; AFM
Documents
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