Biomod/2013/Komaba/Experiment: Difference between revisions
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<h2> <span class="mw-headline">Cylinder</span></h2> | <h2> <span class="mw-headline">Cylinder</span></h2> | ||
<h2> <span class="mw-headline">Ring</span></h2> | <h2> <span class="mw-headline">Ring</span></h2> | ||
<h2> <span class="mw-headline">Spider</span></ | <h2> <span class="mw-headline">Spider</span></h2> | ||
<h1> <span class="mw-headline">Lab Notes</span></h1> | <h1> <span class="mw-headline">Lab Notes</span></h1> | ||
<p>We conducted our experiments at Fuji Laboratory, Komaba Research Campus. | <p>We conducted our experiments at Fuji Laboratory, Komaba Research Campus. | ||
Revision as of 12:05, 24 October 2013
Home,Project,Design,Experiment,Supplementation,Team,Sponsors
Experiments
Cylinder
Ring
Spider
Lab Notes
We conducted our experiments at Fuji Laboratory, Komaba Research Campus.
September 11th Cylinder and Ring(first ver.)
Staple Mix (Cylinder)
| Amount | Stock | ||
|---|---|---|---|
| Staple | 2.5μL(each) | 50μM | |
| TE buffer | 52.5μL | ||
| Total | 500μL |
Stock of the staples are 50μM.
There were 179 strands for the Cylinder.
Cylinder(10 - equivalent)
| Concentration | Amount | Stock | |
|---|---|---|---|
| Mg2+ | 12.5mM | 1.25μL | 1M |
| TAE10x | 10μL | ||
| M13 | 2.5nM | 6.25μL | 40nM |
| Staple Mix | 25nM | 10μL | 250nM |
| mQ | 72.5μL | ||
| Total | 100μL |
Cylinder(15 - equivalent)
| Concentration | Amount | Stock | |
|---|---|---|---|
| Mg2+ | 12.5mM | 1.25μL | 1M |
| TAE10x | 10μL | ||
| M13 | 2.5nM | 6.25μL | 40nM |
| Staple Mix | 37.5nM | 15μL | 250nM |
| mQ | 67.5μL | ||
| Total | 100μL |
Staple Mix (Ring)
| Amount | Stock | ||
|---|---|---|---|
| Staple | 5μL(each) | 50μM | |
| TE buffer | 100μL | ||
| Total | 500μL |
Stock of the staples are 50μM.
There were 80 strands for the Ring.
Ring(10- equivalent)
| Concentration | Amount | Stock | |
|---|---|---|---|
| Mg2+ | 12.5mM | 1.25μL | 1M |
| TAE10x | 10μL | ||
| M13 | 5nM | 12.5μL | 40nM |
| Staple Mix | 50nM | 10μL | 500nM |
| mQ | 66.25μL | ||
| Total | 100μL |
Ring(15 - equivalent)
| Concentration | Amount | Stock | |
|---|---|---|---|
| Mg2+ | 12.5mM | 1.25μL | 1M |
| TAE10x | 10μL | ||
| M13 | 5nM | 12.5μL | 40nM |
| Staple Mix | 75nM | 15μL | 500nM |
| mQ | 61.25μL | ||
| Total | 100μL |
September 13th Observing Cylinder and Ring(first ver.) by AFM
This is the image of the Ring( 15- equivalent).
Unfortunately, since the AFM was out of control for a while, we couldn't take all all the images of our samples which we made on September 11th.
There might have been artificial mistakes, we decided to make the staple mix from the first.
September 13th Agarose Gel Electrophoresis(Cylinder and Ring)
Since we failed to observe our origami by AFM, we decided to try electrophoresis.
Preparing Gel
| Concentration | Amount | |
|---|---|---|
| Agarose | 1.75%(wt/V) | 0.525g |
| TBE10x | 1x | 3mL |
| Mg2+ | 12.5mM | 375μL |
| mQ | up to 30mL | |
| Total | 30mL |
- Pour the gel into frames.
- Put frames into a refrigerator.
Preparing Sample
- For each sample, we mix 2μL of loading buffer and 10μL of DNA sample.
We prepared seven samples - Cylinder(10 - equivalent), Cylinder(15 - equivalent), Cylinder (Staple mix), M13, Ring (Staple mix), Ring(15 - equivalent), Ring(10 - equivalent). - Put seven samples on the gel and pour TBE buffer into the container.
- Leave 1.5 hours under 50V condition.
Observing
We tried to observe this using SyBR Green 1 Dye, but we couldn't to see anything separated. The condition of the gel might not have been good.
September 19th Cylinder and Ring(first ver.)
We retried creating samples of our first version of Cylinder and Ring under exactly the same condition on September 11th.
September 20th Observing Cylinder and Ring(first ver.) by AFM
- This is the image of the Cylinder.
- This is the image of the Ring.
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As you can see, we succeeded in making the Cylinder but we failed to make the Ring.
After a careful consideration, we found out that the design of the structure was not good. Therefore, we designed a new version of the Ring.
October 15th Ring(third ver.)
Staple
Dilute the oligo to 100μM.
Staple Mix
| Concentration | Amount | Stock | |
|---|---|---|---|
| Staple | 1μM | 2μL | 100μM |
| TE buffer | 176μL | ||
| Total | 200μL |
There were 12 strands for this Ring.
Ring
| Concentration | Amount | Stock | |
|---|---|---|---|
| Mg2+ | 14mM | 0.7μL | 1M |
| TBE 10x | 5μL | ||
| Staple Mix | 2.5μM | 25μL | 5μL |
| mQ | 19.3μL | ||
| Total | 50μL |
Annealing Process
- 90°C~15°C
- -0.1°C/min.
- more than 12 hours
