Biomod/2012/Titech/Nano-Jugglers/Methods
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} </style> </head> <BODY> <div id="biomodlink"> <<a href="http://openwetware.org/wiki/Biomod">BIOMOD</a>|<a href="http://openwetware.org/wiki/Biomod/2012">2012</a>|Titech Nano-Jugglers </div> <div id="header"> <div id="navigation"> <div id="menu"> <ul> <li><a href="http://openwetware.org/wiki/Biomod/2012/Titech/Nano-Jugglers"><br>Home<br><br></a></li> <li><a href="http://openwetware.org/wiki/Biomod/2012/Titech/Nano-Jugglers/Team/Students"><br>Team<br><br></a></li> <li><a href="http://openwetware.org/wiki/Biomod/2012/Titech/Nano-Jugglers/Project"><br>Project<br><br></a></li> <li><a href="http://openwetware.org/wiki/Biomod/2012/Titech/Nano-Jugglers/Results">Results<br>&<br>Methods</a></font></li> <li class="ach"><a href="http://openwetware.org/wiki/Biomod/2012/Titech/Nano-Jugglers/Achievements"><br>Achievements<br><br></a> <li class="sup"><a href="http://openwetware.org/wiki/Biomod/2012/Titech/Nano-Jugglers/Protocols"><br>Suppl. Info.<br><br></a></li> <li class="none"><a href="http://openwetware.org/wiki/Biomod/2012/Titech/Nano-Jugglers/Acknowledgement"><br>Acknowledgements<br><br></a></li> </ul> </div> </div> </div> </BODY> </html>
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We have set 3 goals, that is "Energy production for rail-free movement", "Speeding up", and "Direction control". |
Methods
<html><A href=#TOP title="Vapor deposition"><IMG width="300px" src="http://openwetware.org/images/a/a5/Vapor_deposition.jpg"></A></html> <html><A href=#TOP title="SAM conjugation"><IMG width="300px" src="http://openwetware.org/images/4/48/Thiol_.jpg"></A></html> <html><A href=#TOP title="EDAC conjugation"><IMG width="300px" src="http://openwetware.org/images/3/3d/EDAC_conjugation.jpg"></A></html> <html><A href=#TOP title="DNA design"><IMG width="300px" src="http://openwetware.org/images/b/b2/DNA_design.jpg"></A></html> <html><A href=#TOP title="半白金 1μの動画"><IMG width="300px" src="http://openwetware.org/images/8/80/Damy.png"></A></html> <html><A href=#TOP title=DNA hybrudization><IMG width="300px" src="http://openwetware.org/images/f/f1/DNA_hybrudization.jpg"></A></html>
<html><A href=#TOP title="ハイスピードカメラ"><IMG width="300px" src="http://openwetware.org/images/8/80/Damy.png"></A></html> <html><A href=#TOP><IMG width="300px" src="http://openwetware.org/images/e/ed/%E3%82%AB%E3%82%BF%E3%83%A9%E3%83%BC%E3%82%BC.jpg"></A></html> <html><A href=#TOP title="Azobenzene"><IMG width="300px" src="http://openwetware.org/images/e/ea/Azobenzene.jpg"></A></html>
Vapor deposition of Au and Cr on the polystyrene body
File:Body construction.png 
Vapor deposition is a method for forming a thin metal film on the surface of the substrate. In this project, Gold and Chromium were deposited on the polystyrene beads in order to conjugate specific DNA onto the determined location on the surface.
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1st Deposition
2nd Deposition
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After PVD
- Through twice of physical vapor deposition,
- we conjugate thiol modified DNA onto gold part.
- we conjugate amino modified DNA onto polystyrene part.
- There are no DNA staple strands on chromium hemisphere.
- Therefore, platinum particles can't conjugate onto chromium hemisphere.
- As a result, JET starts to move toward chromium hemisphere heading.
- Through twice of physical vapor deposition,
DNA conjugation
File:Body construction.png 
SAM Self-assembled monolayers (SAM) of molecules are molecular assemblies formed spontaneously on substrate surfaces by chemical adsorption. This chemical reactions can be organized into large ordered domains and it is also possible to cover the substrate with functional end group that has some modification. In addition, few steps and experimental technique are required. SAMs are created by adsorption of head groups onto a substrate followed by a slow organization of tail groups.
First, head groups are absorbed onto substrate over . In this phase, the surface of substrate is patterned with spots by dsorbeate molecules rather than beautiful membranes. Over the period of disordering form, the head groups assemble together on the substrate, while the tail groups assemble far from the substrate. Areas of concentrated molecules nucleate and grow until the surface of the substrate is covered in a single monolayer.
Thiol conjugation onto gold
Thiolated compound and gold substrate are one of the most famous research of SAMs. Thiol modified DNA is conjugate to gold particles by this chemical reaction. Initially, substrate gold are cleaned by water or acid-base buffer in order to washout organic impurities. Gold particles are dissolved in 10mM phosphate buffer pH8, 10μM thiol modified DNA. Then in addition to NaCl, raise up to 0.7M salt concentration gradually over a day.Last,clean up again by proper buffer and dry. This reaction is also use with platinum In order to accomplish our project ,we need platinum-DNA and gold-DNA conjugation, so we use thiol modified DNA.
- >>see Result
EDAC We bond polystyrene-beads with DNA using EDAC. EDAC conjugation is the method to conjugate animo modified DNA onto carboxylated polystyrene surface. By using this method, we can connect platinum to aircraft body part of polystyrene regiospecifically.
EDAC is known as water-soluble carbodiimide (WSC) and is used as coupling agent to form amide bond. EDAC crosslinks nucleic acids to the surfaces, which have carboxy group. In this project, we used this method to conjugate amino modified DNA onto carboxylated polystyrene-beads. EDAC reacts with carboxylated polystyrene-beads to form active intermediate on the body. Then it reacts to amino modified DNA. The beads are coupled by nucleic acid. We divide the body into three parts including polystyrene surface, so it is possible to connect platinum regiospecifically. - >>see Result
DNA Design
We used NUPACK and designed five strands of DNA.
- >>see more Method
Observation of platinum hemisphere behavior in solution of H2O2
DNA hybridization in solution of H2O2
- H2O2 has strong corrosion. There was a risk that DNA strand is denatured by H2O2. So, we had to ensure that the hybridization of DNA is not suffer from H2O2. We guess that if DNA had been destroyed, then molecular mass would decrease in that DNA can't hybridize. Measure the relative magnitude of the molecular weight by electrophoresis, we are able to examaine whether DNA can hybridize or not.
- This time, we have to examine the state of the DNA in the time we need to steer rocket, set the time to soak and concentration of H2O2. We used PAGE electrophoresis to ascertain the stability of DNA duplex in thin H₂O₂ solution 1%~5%. PAGE electrophoresis shows the difference of molecular weight that comes from denaturetion or hybridization in the form of bands.
- >>see Result
Analysis of platinum by High-speed camera
Energy production by using catalase
Dissociation of azobenzene-modified DNA by UV-light irradiation
- >>Result page is here
- The duplex-forming activities of DNA can be photomodulated by incorporation of an azobenzene unit. The duplex is dissociated on isomerizing the trans-azobenzene to the cis form by irradiation with UV light. The duplex is formed again when the cis-azobenzene is converted to the transazobenzene by irradiation with visible light.
- We inserted this function into a bead-body and add the photo-switching function to the body.In brief, so that we can detached the engines by irradiation with UV light, we attached a part of engines to the body with DNA duplex incorporating azobenzene unit.
- >>Result page is here
DNA designDesign
Image1 in detail
- 5’ -(HS)-AATxACxCCxAGxCC-3’ (x=azobenzene)
1.We incorporated 4 azobenzene units into 11 bases DNA so that we can quickly dissociated DNA duplex.
2.5’ end of this DNA was modified by a thiol group(HS-) for forming Au-thiol bond with Au covered bead body.
Image2 in detail
- 5’-(HS)-GGCTGGGTATT-3’
1.This 11 bases DNA are a complementary sequence of DNAⅰ(photoresponsive DNA).
2.5’ end of this DNA is modified by a thiol group(HS-) for forming Pt-thiol bond with Pt.
Ascertain the photo-switching system
- Spectrophotometer
- We measured DNA absorbance and ascertain the duplex-forming and duplex-dissociation activities of DNAⅰ and ⅱ. As the ordered regions of stacked base pairs in the DNA duplex are dissociated, the UV absorbance increases. This difference in absorbance between the duplex and single strand state is the result of nearest neighbor base pair interactions. In other words, when the DNA is in the duplex state, interactions between base pairs decrease the UV absorbance relative to single strands. When the DNA is in the single strand state the interactions are much weaker,due to the decreased proximity, and the UV absorbance is higher than the duplex state[image3].
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