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| ==Week 1== | | <div class="tabs-blue"> |
| ===06.12.06: Mon.=== | | <ul> |
| ====Folding DNA Nanostructures====
| | <li id="current">[[IGEM:Harvard/2006/vlau|Contact Info.]]</li> |
| 1. Working Stocks
| | <li>[[IGEM:Harvard/2006/vlau/Lab Notebook|Lab Notebook]]</li> |
| 44 nM scaffold (20 microL)
| | <li>[[IGEM:Harvard/2006/vlau/Presentations|Presentations]]</li> |
| 0.99 microM of each oligo
| | <li>[[IGEM:Harvard/2006/vlau/Protocols|Protocols]]</li> |
| 2. Protocol
| | </ul> |
| - goal: 10 nM scaffold w/ 100 nM each oligo, total rxn vol of 20 microL
| | </div> |
| - calculations:
| | <br style="clear:both"> |
| scaffold = (10 nM)/(44 nM) * 20 microL = 4.5 microL
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| oligos = (100 nM)/(990 nM) * 20 microL = 2 microL
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| - reaction mixture:
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| 4.5 microL p7308 scaffold
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| 2 microL oligos
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| 2 microL 10x folding buffer (500 mM HEPES ph 7.5, 500 mM NaCl, 100 mM MgCl2)
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| 11.5 microL dH2O
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| - annealing times:
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| 90 dC, 5'
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| 65 dC, 20'
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| 55 dC, 20'
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| 45 dC, 20'
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| 37 dC, 30'
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| - neg. controls: 1 replacing oligos w/ water, 1 replacing scaffold w/ water
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| ====Transformation====
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| 1. Plasmids
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| R0010: lac operon promoter
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| E7104: T7 promoter + GFP
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| E0241: GFP
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| 2. Protocol
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| - let chemically competent OneShot Top10 cells thaw on ice
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| - added appropriate DNA to cells and tapped gently to mix
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| - let sit on ice for 20 min
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| - heat shocked @ 42 dC for 30"
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| - let cool on ice for 2 min
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| - added 200 microL SOC media
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| - shook @ 37 dC for 1 hr
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| - pipetted and spread onto agar plates treated w/ ?
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| - incubated @ 37 dC overnight agar side-up
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| | 335 Kirkland House Mail Center |
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| | vlau@fas.harvard.edu | vhtlau@gmail.com |
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| | [[IGEM:Harvard/2006]] |
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| [[June 12 (Monday)]] | | [[IGEM:Harvard/2006/DNA nanostructures]] |
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| [[June 13 (Tuesday)]]
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| [[June 14 (Wednesday)]]
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| [[June 15 (Thursday)]]
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