Name: design and synthesis of a reconfigurable dna accordion rack
Uploaded: 2018-08-15T15:00:00
Description: Watch this Scientific Journal Video about Design and Synthesis of a Reconfigurable DNA Accordion Rack at JoVE.com

This research was partially supported by the Global Research Development Center Program through the National Research Foundation of Korea(NRF) funded by the Ministry of Science and ICT (MSIT) (2015K1A4A3047345) and Nano&#183;Material Technology Development Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT (MSIT) (2012M3A7A9671610). The Institute of Engineering Research at Seoul National University provided research facilities for this work. Authors acknowledge gratitude towards Tae-Young Yoon (Biological Sciences, Seoul National University) regarding the fluorescence spectroscopy for the FRET analysis.

1. Design of the 6 by 6 DNA Accordion Rack with caDNAno14
<ol>
	<li>Download and install caDNAno 2.0 software14 to design a DNA accordion rack (caDNAno 2.5 is also available on https://github.com/cadnano/cadnano2.5). Open caDNAno14 and click the Square Tool to add a new part with a square lattice.</li>
	<li>Number each beam of the accordion rack and draw on the left lattice panel of the caDNAno14 (<strong cl...

<ol>
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This protocol introduces the entire process from design, simulation, synthesis, and analysis of the basic 2D DNA accordion rack. The modified design and simulation rules have been described because the design rule differs from that of standard DNA origami, in that the DNA accordion rack has additional nucleotides at the crossovers for flexibility14,15. From this, we expect that the protocol can accelerate various researches using DNA accordion racks. In addition,...

Mechanical systems based on DNA nanostructures or DNA nanomachines1,2,3,4,5 are unique because they produce complex nanoscale motion in 2D and 3D in the nanometer to &#229;ngstr&#246;m resolution, according to various biomolecular stimuli2,3,6. By attaching functional materials on these structures and controlling their positions, these structures can be applied to various areas. For example, DNA nanomachines have been proposed for a molecular reactor7, drug delivery8, and nanoplasmonic systems9,10.
Previously, we introduced the reconfigurable DNA accordion rack, which can manipulate a 2D or 3D nanoscale network of elements11 (Figure 1A). Unlike other DNA nanomachines that only control a few elements, the platform can collectively manipulate periodically arrayed 2D or 3D elements into various stages. We anticipate that a programmable chemical and biological reaction network or a molecular computing system can be built from our system, by increasing the number of controllable elements. The DNA accordion rack is a structure, in which the network of multiple DNA beams is connected to joints composed of single-stranded DNA (Figure 1B). The accordion rack generated by the DNA beams is reconfigured by the DNA locks, which hybridize to the sticky part of beams and change the angle between the beams according to the length of the bridging part of the locks (locked state). In addition, multi-step reconfiguration is demonstrated by adding new locks after formation of the free state by detaching DNA locks through toehold-based strand displacement12,13.
In this protocol, we describe the entire design and synthesis process of the reconfigurable DNA accordion rack. The protocol includes design, simulation, wet-lab experiments, and analysis for the synthesis of the DNA accordion rack of 6 by 6 meshes and a reconfiguration of these. The structure covered in the protocol is the basic model of the previous research11 and is 65 nm by 65 nm in size, consisting of 14 beams. In terms of the design and simulation, the structural design of the accordion rack is different from conventional DNA origami14,15 (i.e., tightly packed). Therefore, the design rule and molecular simulation have been modified from traditional methods. To demonstrate, we show the design technique using the modified approach of caDNAno14 and the simulation of the accordion rack using oxDNA16,17 with additional scripts. Finally, both protocols of TEM and FRET for analysis of configured accordion rack structures are described.

<table>
	<tbody>
		<tr>
			<td>M13mp18 Single-stranded DNA</td>
			<td>NEB</td>
			<td>N4040s</td>
			<td></td>
		</tr>
		<tr>
			<td>1M MgCl2 Solution</td>
			<td>Biosesang</td>
			<td>M2001</td>
			<td></td>
		</tr>
		<tr>
			<td>Tris-EDTA buffer</td>
			<td>Biosesang</td>
			<td>T2142</td>
			<td></td>
		</tr>
		<tr>
			<td>Nuclease-Free Water</td>
			<td>Qiagen</td>
			<td>129114</td>
			<td></td>
		</tr>
		<tr>
			<td>5M Sodium Chloride solution</td>
			<td>Biosesang</td>
			<td>s2007</td>
			<td></td>
		</tr>
		<tr>
			<td>PEG 8000</td>
			<td>Sigma Aldrich</td>
			<td>1546605</td>
			<td></td>
		</tr>
		<tr>
			<td>10N NaOH</td>
			<td>Biosesang</td>
			<td>S2038</td>
			<td></td>
		</tr>
		<tr>
			<td>Uranyl formate</td>
			<td>Thomas Science</td>
			<td>C993L42</td>
			<td></td>
		</tr>
		<tr>
			<td>Thermal cycler C1000</td>
			<td>Biorad</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Nanodropic 2000</td>
			<td>Thermo Fisher Scientific</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>TEM (LIBRA 120)&#160;&#160;</td>
			<td>Carl Zeiss</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Fluorometer Enspire 2300</td>
			<td>Perkin-Elmer</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Centrifuge</td>
			<td>Labogene</td>
			<td>LZ-1580</td>
			<td></td>
		</tr>
	</tbody>
</table>

design and synthesis of a reconfigurable dna accordion rack

DNA nanostructure-based mechanical systems or DNA nanomachines, which produce complex nanoscale motion in 2D and 3D in the nanometer to &#229;ngstr&#246;m resolution, show great potential in various fields of nanotechnology such as the molecular reactors, drug delivery, and nanoplasmonic systems. The reconfigurable DNA accordion rack, which can collectively manipulate a 2D or 3D nanoscale network of elements, in multiple stages in response to the DNA inputs, is described. The platform has potential to increase the number of elements that DNA nanomachines can control from a few elements to a network scale with multiple stages of reconfiguration.
In this protocol, we describe the entire experimental process of the reconfigurable DNA accordion rack of 6 by 6 meshes. The protocol includes a design rule and simulation procedure of the structures and a wet-lab experiment for synthesis and reconfiguration. In addition, analysis of the structure using TEM (transmission electron microscopy) and FRET (fluorescence resonance energy transfer) is included in the protocol. The novel design and simulation methods covered in this protocol will assist researchers to use the DNA accordion rack for further applications.

The designed 6 by 6 DNA accordion rack is simulated from the oxDNA16,17 and the results are shown in Figure 6. From the simulation result, it was confirmed that the intended structure is formed without distortion of the structure.
The TEM images in Figure 7 are images of configured structures with a lock length ...

Watch this Scientific Journal Video about Design and Synthesis of a Reconfigurable DNA Accordion Rack at JoVE.com

Design and Synthesis of a Reconfigurable DNA Accordion Rack

We describe the detailed protocol for design, simulation, wet-lab experiments, and analysis for a reconfigurable DNA accordion rack of 6 by 6 meshes.

DNA nanostructure-based mechanical systems or DNA nanomachines, which produce complex nanoscale motion in 2D and 3D in the nanometer to &#229;ngstr&#246;m ...

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