How to build containers with scaffolded DNA origami

• Helpful skills
	• Ability to use Adobe Illustrator
	• Ability to program in Python

• Exercise #1: Build a curved sheet using a honeycomb lattice constraint
	• Draw an arbitrary path on a honeycomb lattice
		• Label positive (moving away) and minus (returning back) strands
		• Start on a positive strand
		• Can make enclosed structures
	• Draw staple-strand crossovers on the strand diagram
		• Draw the crossovers in the middle of the 14bp subzones
		• Use position guides to determine the proper placement of crossovers
	• Define staple oligos based on crossovers
		• 42bp per oligo
		• Oligos begin on the subzone marker lines
	• Understand the physical basis for the position guides
		• Draw three connected circles that form a 120-degree angle
		• Looking down the barrels of the helices, strands go clockwise away from the viewer and counter-clockwise towards the viewer
	• Extensions
		• Use fingers (= pairs of positive and negative strands) of varying lengths
		• Create holes
		• Connect rigid domains with flexible linkers (e.g. wireframe tetrahedron with twelve-stranded struts)

• Exercise #2: Write Python program to build the object
	• Coding steps
		• Generate token pointer array representations of staple oligos
			• Initialize 2D token pointer array (7 bases per token)
			• Rerout token pointers at oligo crossover points
			• Generate an array of token pointer paths that covers all tokens a single time each
			• Break down these paths into six-token oligo token pointer arrays
		• Generate token sequence array
			• Break scaffold into virtual strands
			• Break virtual strands into tokens
		• Generate oligo sequences
			• Use oligo token pointer arrays and token sequence array
	• Checking for errors
		• Check to see that each token is represented once and only once in the oligo token pointer arrays

• Honeycomb constraints
	• 42bp between colinear crossovers
	• 120-degree angle between any three helices (honeycomb lattice constraint)
	• Most oligos are 42mers
	• Oligos begin in the middle of 14bp subzones

• Protocol for honeycomb-constrained scaffolded DNA origami
	• Step 1: Draw out honeycomb diagram in Illustrator
	• Step 2: Draw out strand diagram in Illustrator
		• Incorporate proper oligo and scaffold crossovers 
	• Step 3: Write code
	• Step 4: Check for errors
	• Step 5: Order oligos
	• Step 6: Mix DNA together, fold
	• Step 7: Native agarose gel analysis
	• Step 8: Image using electron microscopy and/or atomic-force microscopy