Biomod/2012/Harvard/BioDesign/introduction: Difference between revisions

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But this gave rise to another problem. To use L-DNA as the basis for any structure, we would have to use hundreds of unique L-DNA SSTs to provide the specificity that the SST technique requires (or the strands won't fall into the pre-determined place). Unfortunately, the cost of synthesis rises with the number of unique strands and with L-DNA already more expensive to synthesize, the cost of making any L-DNA SST structure is far too high to pursue development with a standard SST approach.
But this gave rise to another problem. To use L-DNA as the basis for any structure, we would have to use hundreds of unique L-DNA SSTs to provide the specificity that the SST technique requires (or the strands won't fall into the pre-determined place). Unfortunately, the cost of synthesis rises with the number of unique strands and with L-DNA already more expensive to synthesize, the cost of making any L-DNA SST structure is far too high to pursue development with a standard SST approach.
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To solve this price issue, we needed to limit the number of unique strands.  Our team came up focused on ways to reduce the number of unique strands: we got it down to just two.  
To solve this price issue, we needed to limit the number of unique strands.  Our team came up focused on ways to reduce the number of unique strands: we got it down to just two.  
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The solution involved more than just using two sets of strands, as that would create indeterminate structures([http://www.sciencemag.org/content/321/5890/824 Yin et al. 2008]).  We adapted a method of "templating", using the unique D-DNA strands to form a template structure first, to attach L-DNA onto and control the L-DNA layer's shape.
The solution involved more than just using two sets of strands, as that would create indeterminate structures([http://www.sciencemag.org/content/321/5890/824 Yin et al. 2008]).  We adapted a method of "templating", using the unique D-DNA strands to form a template structure first, to attach L-DNA onto and control the L-DNA layer's shape.
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By attaching one L-DNA SST type first, which binds to the template, we can then fill in the spaces with the complimentary second L-DNA SST.  This will avoid any infinite tessellation or uncontrollable growth.  
By attaching one L-DNA SST type first, which binds to the template, we can then fill in the spaces with the complimentary second L-DNA SST.  This will avoid any infinite tessellation or uncontrollable growth.  
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[[Image:L-DNA_A_and_B_template.png]]  
[[Image:L-DNA_A_and_B_template.png]]  
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The result of our project is a method that balances self-assembly with nuclease resistance, specifically L-DNA nanostructure templated onto a D-DNA sheet.  With this technique, we can create a size-determined L-DNA sheet - the first step towards our L-DNA nanorobot.
The result of our project is a method that balances self-assembly with nuclease resistance, specifically L-DNA nanostructure templated onto a D-DNA sheet.  With this technique, we can create a size-determined L-DNA sheet - the first step towards our L-DNA nanorobot.
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Revision as of 23:20, 27 October 2012

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Introduction


Background

In February 2012, Shawn M. Douglas, Ido Bachelet, and George M. Church at Harvard University demonstrated the use of DNA origami to create a drug-delivering nanorobot. This device carries with it remarkable advantages including cell-specificity, biocompatibility, and the ability to protect fragile, toxic, or previously undeliverable drugs.

Problem

While an exciting advancement in drug delivery, the DNA nanorobot may be less structurally stable when inside the body: DNA can be degraded by nucleases, enzymes found in the body which cut and digest DNA.


Professor Paul Rothemund, a well-known pioneer in DNA nanotechnology, noted in an accompanying Nature article that "[i]f these sorts of problems can be solved, then the nanorobots have a chance at becoming real therapeutics."

Inspired by the DNA nanorobot's potential as such a future therapeutic, our team decided to tackle the issue of nuclease resistance.

Solution

We focused on one possible method to create nuclease resistant structures: by using a different material. We decided to work with L-DNA as our enzyme-resistant nanomaterial. L-DNA, the mirror image of D-DNA, cannot be recognized and degraded by the chiral nucleases found naturally in our body.


While exciting, the use of L-DNA alone is not an immediate solution. The DNA nanorobot, made with the DNA origami technique, requires the use of a long phage-derived scaffold strand (7,308 base pairs). Because L-DNA is completely synthetic, we cannot use a phage to take base this scaffold off of. And to synthesize L-DNA of this length would be extremely difficult as the error rate will build up with such a long sequence.

Confronted this issue, our team decided to utilize the new technique of Single-Stranded Tiles (SSTs), a different method of creating nano structures. In short (no pun intended), the SST approach consists of shorter, modular strands, rather than stapling together one long strand. The idea was created by Peng Yin in 2008 and explored by Bryan Wei et. al. in 2012.

Before we continue, here's a quick video of how the SST technique works, courtesy of the Wyss Institute.

<html> <center> <iframe src="http://player.vimeo.com/video/42849360?title=0&amp;byline=0&amp;portrait=0&amp;badge=0" width="500" height="281" frameborder="0" webkitAllowFullScreen mozallowfullscreen allowFullScreen></iframe> </center> <h5><p><a href="http://vimeo.com/42849360">How Single-Stranded Tiles (SSTs) Work</a> direct link.</p></h5>

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Unlike DNA origami, the SST method only requires short, 42-mer DNA strands, allowing us to use L-DNA.


But this gave rise to another problem. To use L-DNA as the basis for any structure, we would have to use hundreds of unique L-DNA SSTs to provide the specificity that the SST technique requires (or the strands won't fall into the pre-determined place). Unfortunately, the cost of synthesis rises with the number of unique strands and with L-DNA already more expensive to synthesize, the cost of making any L-DNA SST structure is far too high to pursue development with a standard SST approach.


To solve this price issue, we needed to limit the number of unique strands. Our team came up focused on ways to reduce the number of unique strands: we got it down to just two.


The solution involved more than just using two sets of strands, as that would create indeterminate structures(Yin et al. 2008). We adapted a method of "templating", using the unique D-DNA strands to form a template structure first, to attach L-DNA onto and control the L-DNA layer's shape.


By attaching one L-DNA SST type first, which binds to the template, we can then fill in the spaces with the complimentary second L-DNA SST. This will avoid any infinite tessellation or uncontrollable growth.


The result of our project is a method that balances self-assembly with nuclease resistance, specifically L-DNA nanostructure templated onto a D-DNA sheet. With this technique, we can create a size-determined L-DNA sheet - the first step towards our L-DNA nanorobot.

Project Goals

To consider our project a success, our team listed potential project milestones:

  1. Create a modified D-DNA template
  2. Bind one set of L-DNA strands to the template
  3. Purify the scaffolds away from the excess unbound L-DNA SST Type A
  4. Bind L-DNA SST Type B onto purified scaffolds
  5. Document the cheap, nuclease-resistant L-DNA nanostructures made using SSTs and an elegant scaffolding method.
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