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Designing, Building, and Using DNA Nanoboxes: - Inside-Outside Specific Protection of Sites -

Designing, Building, and Using DNA Nanoboxes: - Inside-Outside Specific Protection of Sites -. Tiffany Chan - Harvard International Genetically Engineered Machines (iGEM) Team. 1. Intro to DNA Nanoboxes. WHAT : Building DNA nanostructures that act as boxes -

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Designing, Building, and Using DNA Nanoboxes: - Inside-Outside Specific Protection of Sites -

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  1. Designing, Building, and Using DNA Nanoboxes: -Inside-Outside Specific Protection of Sites- Tiffany Chan - Harvard International Genetically Engineered Machines (iGEM) Team

  2. 1. Intro to DNA Nanoboxes • WHAT: Building DNA nanostructures that act as boxes - • WHY: Boxes as drug-delivery vehicles - • HOW: • Staple-oligonucleotides + single-stranded scaffold DNA = DNA origami! - • Honeycomb lattice system

  3. HOW: DNA Nanostructures

  4. 2. The Process: Designing and Building a DNA Nanobox • Coming up with ideas • lllustrator diagrams • ASCII diagrams for program inputs • Schematics of the oligo-scaffold hybridizations • Running the program to get a list of oligos • Organizing oligos • Ordering oligos (with any necessary modifications) • Making up oligo solutions • Folding the oligos and scaffold to make the structure • Imaging through gel shifts and electron micrography

  5. A – C: Ideas and Pictures Double-ply hexagonal barrel, smaller so no lids necessary? Single-ply rectangular barrel, lids extending from barrel? Single-ply hexagonal barrel, two lids on the same scaffold? Double-ply hexagonal barrel, two lids on a separate scaffold? A. Coming up with ideas B. lllustrator diagrams C. ASCII diagrams for program inputs

  6. 3. Using DNA Nanoboxes: • GOAL: Make sure they’re actually boxes! • A box defines a separate “inside” and “outside” - • SOLUTION: Protection assays • Protect what on the inside? • Thrombin (stuck to thrombin DNA-aptamers built into the oligos • Streptavidin (stuck to the biotin molecules on the oligos ordered that way) • Oligo-ligand (a double-stranded DNA segment made from a ssDNA staple-end and a ssDNA added to the box mixture after folding) • From what on the outside? • Proteases (trypsin and proteinase K) • Restriction enzyme (AscIII)

  7. 4. Future Plans • More designs • More protection assays • More electron microscopy snakes on a grid

  8. THE END!

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