IGEM:Harvard/2006/DNA nanostructures/Notebook

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==Daily notebook==
<calendar>
<calendar>
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view=threemonths
view=threemonths
format=%name/%year-%month-%day
format=%name/%year-%month-%day
-
weekstart=14
+
weekstart=7
</calendar>
</calendar>
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<calendar>
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name=IGEM:Harvard/2006/DNA_nanostructures/Notebook
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date=2006/10/01
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view=threemonths
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format=%name/%year-%month-%day
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weekstart=7
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</calendar>
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==General Notes==
 +
*'''Stock Oligos''': reconstituted (50uM for non-biotinylated oligos, 200uM for biotinylated) oligos in wells or tubes
 +
*'''Pre-working Stocks''': mixtures of 10uM of each oligo needed for a "class" of oligos (ex. core, barrel, lid1, lid2); for non-biot oligos, this requires 10uL of each of the necessary stock oligos, for biot-oligos, this requires 2.5uL of stock oligos + 7.5uL dH2O
 +
*'''Working Stocks''': mixtures of however-many-oligos-are-in-each-class uL (ex. if there are 94 oligos in the core class of 3.2's design, use 94uL of the core pre-working stock), diluted to a total of 200uL of solution (each individual oligo will be at 250nM)
 +
 +
==Brainstorming and future projects==
 +
 +
===Thrombin-aptamer experiments===
 +
 +
====Questions / procedures====
 +
* what percent gel? 10% to 20% polyacrylamide gels, no SDS (but would make for a good control)
 +
* what incubation conditions?
 +
* how much protein and DNA? protein at 1 {{um}}, DNA at 2 {{um}}
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* Coomassie stain
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 +
====Experiments====
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{| {{table}}
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| align="center" style="background:#f0f0f0;"|number
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| align="center" style="background:#f0f0f0;"|thrombin
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| align="center" style="background:#f0f0f0;"|aptamer
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| align="center" style="background:#f0f0f0;"|nanotube
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| align="center" style="background:#f0f0f0;"|DNA-stained prediction
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| align="center" style="background:#f0f0f0;"|protein-stained prediction
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|-
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|0||-||-||-||no bands||no bands
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|-
 +
|1||-||-||+||slow band (nanotube)||no bands
 +
|-
 +
|2||-||+||-||fast band (aptamer)||no bands
 +
|-
 +
|3||-||+||+||slow band (aptamer-nanotube), traces of fast band (aptamer)||no bands
 +
|-
 +
|4||+||-||-||no bands||fast band (thrombin)
 +
|-
 +
|5||+||-||+||slow band (nanotube)||fast band (thrombin)
 +
|-
 +
|6||+||+||-||medium band (aptamer-thrombin), fast band (aptamer)||medium band (aptamer-thrombin), traces of fast band (thrombin)
 +
|-
 +
|7||+||+||+||very slow band (thrombin-aptamer-nanotube), slow band (aptamer-nantotube), traces of fast band (aptamer)||very slow band (thrombin-aptamer-nanotube), medium band (aptamer-thrombin), traces of fast band (thrombin)
 +
|-
 +
|}
 +
 +
====Buffers====
 +
* Macaya's and Bock's selection buffer: 20 mM Tris-acetate, pH 7.4, 140 mM NaCl, 5 mM KCl, 1 mM CaCl<sub>2</sub>, 1 mM MgCl<sub>2</sub>
 +
* Liu's incubation buffer: 40 mM Tris, 20 mM CH<sub>3</sub>COOH, 2mM EDTA, 12.5 mM (CH<sub>3</sub>COO)<sub>2</sub>Mg, pH 8.0
 +
* Liu's PAGE buffer: 1x TAE/Mg<sup>2+</sup>

Current revision


Contents

Daily notebook

June
SMTWTFS
123
45678910
11121314151617
18192021222324
252627282930
July
SMTWTFS
1
2345678
9101112131415
16171819202122
23242526272829
3031
August
SMTWTFS
12345
6789101112
13141516171819
20212223242526
2728293031


September
SMTWTFS
12
3456789
10111213141516
17181920212223
24252627282930
October
SMTWTFS
1234567
891011121314
15161718192021
22232425262728
293031
November
SMTWTFS
1234
567891011
12131415161718
19202122232425
2627282930


General Notes

  • Stock Oligos: reconstituted (50uM for non-biotinylated oligos, 200uM for biotinylated) oligos in wells or tubes
  • Pre-working Stocks: mixtures of 10uM of each oligo needed for a "class" of oligos (ex. core, barrel, lid1, lid2); for non-biot oligos, this requires 10uL of each of the necessary stock oligos, for biot-oligos, this requires 2.5uL of stock oligos + 7.5uL dH2O
  • Working Stocks: mixtures of however-many-oligos-are-in-each-class uL (ex. if there are 94 oligos in the core class of 3.2's design, use 94uL of the core pre-working stock), diluted to a total of 200uL of solution (each individual oligo will be at 250nM)

Brainstorming and future projects

Thrombin-aptamer experiments

Questions / procedures

  • what percent gel? 10% to 20% polyacrylamide gels, no SDS (but would make for a good control)
  • what incubation conditions?
  • how much protein and DNA? protein at 1 μM, DNA at 2 μM
  • Coomassie stain

Experiments

number thrombin aptamer nanotube DNA-stained prediction protein-stained prediction
0---no bandsno bands
1--+slow band (nanotube)no bands
2-+-fast band (aptamer)no bands
3-++slow band (aptamer-nanotube), traces of fast band (aptamer)no bands
4+--no bandsfast band (thrombin)
5+-+slow band (nanotube)fast band (thrombin)
6++-medium band (aptamer-thrombin), fast band (aptamer)medium band (aptamer-thrombin), traces of fast band (thrombin)
7+++very slow band (thrombin-aptamer-nanotube), slow band (aptamer-nantotube), traces of fast band (aptamer)very slow band (thrombin-aptamer-nanotube), medium band (aptamer-thrombin), traces of fast band (thrombin)

Buffers

  • Macaya's and Bock's selection buffer: 20 mM Tris-acetate, pH 7.4, 140 mM NaCl, 5 mM KCl, 1 mM CaCl2, 1 mM MgCl2
  • Liu's incubation buffer: 40 mM Tris, 20 mM CH3COOH, 2mM EDTA, 12.5 mM (CH3COO)2Mg, pH 8.0
  • Liu's PAGE buffer: 1x TAE/Mg2+
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