Biomod/2011/TeamJapan/Sendai/Strategy
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<ul id="verticalmenu" class="glossymenu"> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai">Home</a></li> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Strategy">Strategy</a></li> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Design">Design</a></li> <li><a href="#">Experiments</a>
<ul> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Results/Electrophoresis">Electrophoresis</a> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Results/Atomic_Force_Microscope">AFM</a> </ul>
</li> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Computational_design/Simulation" >Simulation</a></li> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Notes">Notes</a></li> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Team">Team</a></li> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Resources">Resources</a></li> <li><a href="http://openwetware.org/wiki/Biomod/2011/TeamJapan/Sendai/Sitemap">Sitemap</a></li>
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Project summary
Designing a molecular robot is one of the most interesting and challenging targets in biomolecular design. This year, two teams from Japan and one from Denmark propose the first “molecular race” over a defined track made with DNA origami. We are now making a molecular robot for the race.
Our molecular robot and its mechanism for movement are based on the molecular spider developed by Lund et al. (Nature, 2010). In the original design, the spider body consisted of the streptavidin protein, and three DNA-based legs are attached to it. The walking movement of the spider is random, thus the robot must be controlled by means of the patterned course on the Origami.
To win the race, we want to substantially improve the robot performance. For this purpose, we make the whole structure of our robot with DNA, which allows us to design arbitrary geometry of the body. Also, we can assign different base sequence to each leg and scaffold on the field. These new parameters give us freedom to optimize our robot design.
We have been developing a stochastic dynamics simulation model in order to evaluate the movement of different types of molecular robots, searching for the optimal design. In the final report, we will show our optimal design and the experimental results including walking motion of the robot captured by a video-rate AFM.
Molecular Robot Race
Rules
A Molecular robot's task is to move from the start region to the goal region , as fast as possible.
The field is placed on a cleaved mica surface using counter ion method.
No restriction is defined for the solution environment, as long as the field and the movement of robot are observable by fast scanning AFM.
No restriction is defined as a material of the robot.
Our strategy
We want to get to the goal more faster.
Now, what should you do make a robot arrive at the goal more quickly?
First, we thought of a person to a model.
1 enlarging steps
2 reducing useless motion
3 moving legs quickly
Second,in molecular world
1 enlarging a body or legs
2 controlling the random motion
3 cutting substrate faster
As a result, to be real.
1 making a larger body than streptavidin by DNA origami method
2 using some kind of legs to restrain random walking
3 improving substrates to increase the efficiency of cutting system
Project
Designing the molecular robot
The body of our molecular robot has the shape of a triangular prism. Previous to BIOMOD2011, we have never made any DNA structure. Therefore, we thought it difficult to make a 3D structure and more than that to view it.
One of the problems in visualizing this kind of structure arise from the fact that an AFM observation is done from the top of the sample surface. So, in the case of the triangular prism we may not probably distinguish whether what we detect is our desire structure or not.
Under the previous circumstances, we decided to make a 2D structure: a development view taken from the structure of a triangular prism. Figure1 and figure 2 show our 2D structure design using caDNAno and its assembled view, respectively. We planned that if both ends of the 2D structure are connected by double strands between the green and red staple (Figure 3), our proposed structure is complete.
Figure 3. caDNAno design: Schematic design of M13mp18 (thin line colored sky-blue) and staples
Our robot moving style
We considered our robot to move along a specific direction just by rolling. Our initial plan consisted in attaching the two same legs, of three different kinds, to each edge of the triangular prism. We got a better performance just by solely using one kind of legs instead of using three in the simulation. So, we decided to attach two legs of one kind of legs to each peak of triangular prism.
Cutting DNA
For producing the robot body we used the viral M13mp18 DNA single strand (M13) as scaffold.
But we only used 1,108 bases of 7,249 bases ([1]), then having a leftover.
In this situation we thought about cutting the part of M13 that we needed.
Our first attempt was to extract the necessary part of M13 to reproduce M13 with polymerase chain reaction.
But we failed.
So we changed our method for cutting the M13 with restriction enzyme.
Therefore, we found that the restriction enzyme method is an easy way to get and can cut near 1,108 bases, we later checked by electrophoresis whether the M13 sequence was properly cut.
3D DNA nanostructure
- Electrophoresis
First, we carried out electrophoresis (EP) in order to check whether the structure was made. We did EP only for the M13 and did annealing for the sample mix of M13 and staples, and analyzed the length difference between the bands. We used agarose as a gel.
- Atomic force microscope
In this stage that we concluded making our structure for EP, we observed by atomic force microscope (AFM) the sample after annealing. First, we observed the 2D structure. Second, as this was success we proceeded to check the 3D structure. As mentioned in section "Designing the molecular robot", the reason was that we judged after observation of 2D structure, having calculation of 3D structure's view, we observe 3D structure were easier than don't having.
Preparation of our structure and field from solution
When we carried out annealing, we added a more quantity of staples than M13s. Therefore, we tried to get rid of the over staples from sample. Because of the excess of staples, we can hardly distinguish between body structure and field.
We have three methods to separate the excess staples from sample:
These methods could remove the excess staples, however it is not sure to take safely the body structure from solution.
