Folding DNA to Create Nanoscale Shapes and Patterns Paul W. K. Rothemund Nature, V440, 297-302, 2006 - PowerPoint PPT Presentation

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Folding DNA to Create Nanoscale Shapes and Patterns Paul W. K. Rothemund Nature, V440, 297-302, 2006

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Building DNA patterns and shapes with a long ssDNA and a bunch of ... DNA nonostructure patterning may be used as templates for programmed molecular arrays ... – PowerPoint PPT presentation

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Title: Folding DNA to Create Nanoscale Shapes and Patterns Paul W. K. Rothemund Nature, V440, 297-302, 2006


1
Folding DNA to Create Nanoscale Shapes and
PatternsPaul W. K. RothemundNature, V440,
297-302, 2006
  • Sunmin Ahn
  • Journal Club Presentation
  • October 23, 2006

2
Outline
  • Introduction
  • Review of DNA structure
  • Designing DNA origami
  • Folding with viral genome
  • Patterning
  • Conclusion

3
Introduction
  • Parallel synthesis of nanostructures
  • Building DNA patterns and shapes with a long
    ssDNA and a bunch of staple strands
  • One pot self assembly

4
DNA Structure
5
Designing Pattern
- Manual design
1. Generation of block diagram
2. Generation of a folding path - raster fill
pattern must be hand designed
6
Designing Pattern
- Computer aided
3. Generation of a first pass design - raster
fill pattern must be hand designed - no bases
left unpaired - single phosphate from each
backbone occurs in the gap - small angle bending
does not affect the width of DNA origami
7
Designing Pattern
- Computer aided
4. Refinement of the helical domain length - to
minimize strain in design - twist of scaffold
calculated and scaffold x-over strains are
balanced by a single bp change - periodic
x-overs of staples are arranged with glide
symmetry ? minor groove faces alternating
directions in alternating columns
8
Designing Pattern
- Computer aided
5. Breaking and merging of strands - pairs of
adjacent staples are merged to yield fewer,
longer staples - merge patterns are not
unique - staggered merge strengthens seam
9
Designing Pattern
- Computer aided
5. Breaking and merging of strands - rectilinear
merge
10
Folding viral genome
  • Circular genomic DNA from virus M13mp18 chosen as
    a scaffold
  • Naturally ssDNA 7249-nt long
  • For linear scaffold 73-nt region containing 20-bp
    stem hairpin was cut with BsrBI restriction
    enzyme
  • resulting 7167nt long linear strand
  • 100X excess of staples and short (lt25nt)
    remainder strands mixed with scaffold and
    annealed 95ºC to 20ºC in a PCR machine (lt 2
    hours)
  • Samples deposited on mica and imaged with AFM in
    tapping mode

11
Folding viral genome
  • Square
  • linear scaffold
  • 13 well formed
  • 25 rectangular fragments
  • 25 hourglass fragments
  • Rectangle
  • tests bridged seam
  • circular scaffold
  • 90 well formed

1µm scale bars
12
Folding viral genome
  • Star
  • demonstrates certain arbitrary shape
  • linear and circular scaffold
  • 11 and 63 well formed
  • higher of well formed shapes with circular
    scaffold may be due to higher purity of the
    scaffold strand

Circular scaffold
100nm scale bars
Linear scaffold
  • Smiley
  • circular scaffold
  • need not be topological disc
  • 90 well formed
  • narrow structures are difficult to form ?
    provides weak spot

100nm scale bar
13
Folding viral genome
  • Triangle from 3 rectangles
  • single covalent bond holding the scaffold
    together
  • less than 1 well formed
  • stacking

100nm scale bar
  • Triangle built from 3 trapezoids
  • circular scaffold
  • 88 well formed with bridging staples
  • 55 well formed without bridging staples

100nm scale bar
14
Stacking
Interaction between blunt end helices cause
stacking
A
B
  1. Staple strands on the edge may be removed (B)
  2. Addition of 4T hairpin loops (F)
  3. Addition of 4T tails on staples that has ends on
    the edge of the shape (D)

Stacked rectangles
Staple strand on the edge removed
F
C
D
Normal amount of aggregation (Smileys)
Addition of 4T tails
1µm scale bars
15
Defects and Damages
100nm scale bars
16
Stoichiometry
  • In most experiments 100300 fold excess over
    scaffold was used
  • 10 fold excess is safe, but not a fundamental
    requirement
  • 2-fold excess may be used

1µm scale bars
17
Patterning
18
Patterning
Binary patterning 1 3nm above mica
surface 0 1.5nm above mica surface
1µm scale bars
19
Patterning
  • Infinite periodic structures are made using
    extended staples
  • Stoichiometry becomes very important
  • 30 Megadalton structure (individual origami
    4megadalton)

100nm scale bars
20
Difficulties
  • Blunt end stacking
  • Down hairpin loops
  • But mostly AFM imaging!!!

21
What about 2º Structures?
  • Lowest E folds calculated

Strong structure
Weak structure
  • Average -965-37kcal/mole
  • Random 6000 base sequence generated with same
    base composition as M13mp18
  • - Similar 2º structure
  • - Average free E -867 - 13kcal/mole

22
How does it work?
  1. Strand invasion
  2. Excess of staples
  3. Cooperative effects
  4. Designs that doesnt allow staples to bind to
    each other

23
Conclusion
  • Quantitative and statistical analysis
  • Better imaging technique should be implemented
  • DNA nonostructure patterning may be used as
    templates for programmed molecular arrays
  • Protein arrays
  • nanowires
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