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

Denne forskning blev delvist støttet af den globale forskning Development Center Program gennem den nationale forskning fundament af Korea(NRF) finansieret af Ministeriet for videnskab og IKT (MSIT) (2015K1A4A3047345) og Nano· Materielle teknologi Development Program gennem National Research Foundation af Korea (NRF) finansieret af Ministeriet for videnskab og IKT (MSIT) (2012M3A7A9671610). Institute of Engineering Research på Seoul National University forudsat forskningsfaciliteter til dette arbejde. Forfatterne anerkender taknemmelighed mod Tae-unge Yoon (biologiske videnskaber, Seoul National University) vedrørende fluorescens spektroskopi til FRET analyse.

1. design af 6 af 6 DNA harmonika Rack med caDNAno14
<ol><li>Hent og Installer caDNAno 2.0 software14 for at designe et DNA harmonika rack (caDNAno 2,5 er også tilgængelig på https://github.com/cadnano/cadnano2.5). Åbn caDNAno14 og klik på Pladsen værktøj for at tilføje en ny del med et firkantet gitter.</li>
	<li>Nummerere hver stråle af harmonika rack og trække på venstre gitter panel af caDNAno14</su...

<ol>
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Denne protokol indfører hele processen fra design, simulering, syntese og analyse af den grundlæggende 2D DNA harmonika rack. Den modificerede design og simulering regler er blevet beskrevet fordi design reglen adskiller sig fra standard DNA origami, DNA harmonika rack har yderligere nukleotider på delefiltre for fleksibilitet14,15. Fra dette forventer vi, at protokollen kan accelerere forskellige forsker ved hjælp af DNA harmonika stativer. Derudover kan bes...

Mekaniske systemer baseret på DNA-nanostrukturer eller DNA nanomaskiner1,2,3,4,5 er unikke, fordi de producerer komplekse nanoskala bevægelse i 2D og 3D i nanometer til Ångström resolution, ifølge forskellige Biomolekylær stimuli2,3,6. Ved fastgørelse funktionelle materialer på disse strukturer og kontrollere deres positioner, kan disse strukturer anvendes på forskellige områder. For eksempel har DNA nanomaskiner været foreslået for en molekylær reaktor7, drug delivery8og nanoplasmonic systemer9,10.
Tidligere introducerede vi den omkonfigurerbare DNA harmonika rack, som kan manipulere en 2D eller 3D nanoskala netværk af elementer11 (figur 1A). I modsætning til andre DNA nanomaskiner, der kun et par betjeningselementer, kan platformen kollektivt manipulere regelmæssigt klædt 2D eller 3D elementer i forskellige stadier. Vi forventer, at en programmerbar kemiske og biologiske reaktion netværk eller en molekylær computing system kan bygges fra vores system, ved at øge antallet af kontrollerbare elementer. DNA harmonika rack er en struktur, hvor netværk af flere DNA bjælker er tilsluttet samlinger består af enkeltstrenget DNA (figur 1B). Harmonika rack genereret af DNA bjælker er omkonfigureres af DNA-låse, som krydse til den klæbende del af bjælker og ændre vinklen mellem bjælker efter længden af den "passerelle" del af låse (låst tilstand). Derudover er flertrins omkonfiguration påvist ved at tilføje nye låse efter dannelsen af den frie tilstand ved at fjerne DNA låse gennem fodfæste-baserede strand forskydning12,13.
I denne protokol beskriver vi hele design og syntese processen med omkonfigurerbare DNA harmonika rack. Protokollen omfatter design, simulering, våd-lab eksperimenter og analyse for syntesen af DNA harmonika rack af 6 af 6 masker og en rekonfigurering af disse. Strukturen er omfattet af protokollen er den grundlæggende model af den tidligere forskning11 og er 65 nm ved 65 nm i størrelse, bestående af 14 bjælker. Med hensyn til design og simulering er den strukturelle design af harmonika rack forskellige fra konventionelle DNA origami14,15 (dvs. stramt pakket). Derfor er design regel og molekylær simulation blevet ændret fra traditionelle metoder. For at demonstrere, viser vi den design teknik ved hjælp af den modificerede tilgang af caDNAno14 og simulering af harmonika rack med yderligere scripts oxDNA16,17 . Endelig, begge protokoller TEM og FRET for analyse af konfigurerede harmonika rack strukturer er beskrevet.

<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 nanostrukturer-baserede mekaniske systemer eller DNA nanomaskiner, der producerer komplekse nanoskala bevægelse i 2D og 3D i nanometer ångström beslutning, viser stort potentiale inden for forskellige områder af nanoteknologi som de molekylære reaktorer, medicinafgivelse, og nanoplasmonic systemer. Den omkonfigurerbare DNA harmonika rack, som kan kollektivt manipulere en 2D eller 3D nanoskala netværk af elementer, i flere stadier i svar til DNA input, er beskrevet. Platformen har potentiale til at øge antallet af elementer, der DNA nanomaskiner kan styre fra et par elementer til en netværk skala med flere faser af omkonfiguration.
I denne protokol beskriver vi hele eksperimentelle processen med omkonfigurerbare DNA harmonika rack af 6 af 6 masker. Protokollen indeholder en design regel og simulation procedure af strukturerne og en våd-lab eksperiment for syntese og rekonfiguration. Derudover er analyse af strukturen ved hjælp af TEM (transmissions elektron mikroskopi) og FRET (Fluorescens resonans energioverførsel) medtaget i protokollen. De nye design og simulering metoder er omfattet af denne protokol vil hjælpe forskere at bruge DNA harmonika rack for yderligere programmer.

Designet 6 af 6 DNA harmonika rack er simuleret fra oxDNA16,17 , og resultaterne er vist i figur 6. Fra simuleringen resultatet, blev det bekræftet, at den tilsigtede struktur er dannet uden forvrængning af strukturen.
TEM billeder i figur 7 er billeder af konfigurerede strukturer med en lås længde på 2, 8,...

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

Vi beskriver detaljerede protokollen til design, simulering, våd-lab eksperimenter og analyse for en omkonfigurerbare DNA harmonika rack af 6 af 6 masker.

Design og syntese af en omkonfigurerbare DNA harmonika 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 ...

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