Team:Goettingen/Project/Materials

From 2012.igem.org

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==== 5x M9 salts stock solution ====
==== 5x M9 salts stock solution ====
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Stir until dissolved; sterilize by autoclaving at 121°C. <br>
 
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{| class="goetable"
!Amount !! Substance
!Amount !! Substance
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* Stir until dissolved; sterilize by autoclaving at 121°C.
==== CaCl<sub>2</sub> Buffer for competent cells ====
==== CaCl<sub>2</sub> Buffer for competent cells ====
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Stir until dissolved; sterilize by sterile filtration.
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{| class="goetable"
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<html><table width="50%" border="1" bordercolor="#000000" cellpadding="0" cellspacing="2">
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!Amount !! Substance !! Notes
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<tr>
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|-
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  <th scope="col">Amount</th>
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| 7.351 g|| CaCl<sub>2</sub> &times; 7 H<sub>2</sub>O || Alternative 0.1 M CaCl<sub>2</sub>.
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  <th scope="col">Substance</th>
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|-
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  <th scope="col">Notes</th>
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|15% || Glycerol ||
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</tr>
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|-
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<tr>
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|ad 500 ml || ddH<sub>2</sub>O ||
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  <td>7.351 g </td>
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  <td>CaCl<sub>2</sub> &times; 7 H<sub>2</sub>O</td>
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|}
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  <td>Alternative 0.1 M CaCl<sub>2</sub>.</td>
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* Stir until dissolved; sterilize by sterile filtration.
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</tr>
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<tr>
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  <td>15%         </td>
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  <td> Glycerol</td>
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  <td></td>
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</tr>
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  <tr>
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  <td>ad 500 ml</td>
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  <td>ddH<sub>2</sub>O</td>
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  <td></td>
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</tr>
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</table></span></html>
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==== 50x TAE for agarose gels ====
==== 50x TAE for agarose gels ====
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Stir until dissolved. Working concentration is 1x (40 ml of 50x stock ad 2 l ddH<sub>2</sub>O).
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<html><span style="text-indent:20px;"><table width="50%" border="1" bordercolor="#000000" cellpadding="0" cellspacing="2">
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! Amount !! Substance
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  <th scope="col">Amount</th>
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|242 g || Tris base
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  <th scope="col">Substance</th>
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</tr>
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|57.1 m l| Glacial acetic acid
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  <td>242 g</td>
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|18.6 g || EDTA
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  <td>Tris base</td>
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</tr>
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|ad 1000 ml | ddH<sub>2</sub>O
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<tr>
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  <td>57.1 ml        </td>
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|}
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  <td>Glacial acetic acid </td>
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* Stir until dissolved. Working concentration is 1x (40 ml of 50x stock ad 2 l ddH<sub>2</sub>O).
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</tr>
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  <tr>
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  <td>18.6 g</td>
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  <td>EDTA </td>
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</tr>
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  <tr>
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  <td>ad 1000 ml </td>
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  <td>ddH<sub>2</sub>O </td>
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</tr>
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</table></span></html>
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== <i>E. coli</i> ==
== <i>E. coli</i> ==
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Storage of <i>E. coli</i> strains - How to keep <i>E. coli</i> strains.
Storage of <i>E. coli</i> strains - How to keep <i>E. coli</i> strains.
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* Glycerol Stock
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==== Glycerol Stock ====
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** 900 µl over night culture in LB-liquid medium
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The glycerol stock is prepared as follow:
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** 900 µl 50% sterile glycerol
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* 900 µl over night culture in LB-liquid medium
 +
* 900 µl 50% sterile glycerol
Glycerol is extremely viscous. Hence, use cut off tips to pipette. Glycerol stocks were vortexed to ensure uniformly mixture and shock
Glycerol is extremely viscous. Hence, use cut off tips to pipette. Glycerol stocks were vortexed to ensure uniformly mixture and shock

Revision as of 14:49, 22 September 2012

Deutsch  / English 

Contents

Recipes

Media

Recipes for liquid and solid media for bacterial cultivation. Stir until dissolved; sterilize by autoclaving 121°C. Do not forget a magnetic stirrer.

LB-Medium

  • 1% (w/v) bacto tryptone
  • 0.5% (w/v) bacto yeast extract
  • 1% (w/v) NaCl
  • if required: ad 1% (w/v) agar

M9 Swimming Agar

  • 1.25% (v/v) glycerol
  • 20% (v/v) 5x M9 salt stock solution
  • 0.1% (v/v) of CaCl2 - 2H2O stock solution (20 mg/mL)
  • 0.1% (v/v) MgSO4 stock solution (0.12 g/mL)
  • 0.3 % (w/v) of agar

Tryptone swimming agar

  • 1% (w/v) bacto tryptone
  • 0.5% (w/v) NaCl
  • 0.3% (w/v) agar

Antibiotics

Stock solutions and concentrations of antibiotics.

Supplements

  • Aminoacid mix (containing all aminoacids except tryptophane): stocksolution 0.2% (w/v) varying amounts used in swimming assays, view methods and notebook of group 1
  • Caffeine: stocksolution 10 mM 100 µl applied to whatmanpaper in swimming assays, view methods
  • D-Aspartate: stocksolution 10 mM
  • 2-Ethyl-1-Hexanole: stocksolution 5 mM 100 µl applied to whatmanpaper in swimming assays, view methods
  • Geraniol: stocksolution 5 mM 100 µl applied to whatmanpaper in swimming assays, view methods
  • L-Aspartate-4-Benzyl: stocksolution 10 mM 100 µl applied to whatmanpaper in swimming assays, view methods
  • L-aspartate: stocksolution 10 mM varying amounts used in swimming assays, view methods and notebook of group 1
  • Leucin: stocksolution 30.49 mM add to M9-agar: 4 ml (final concentration 0.305 µM)
  • Methionin: stocksolution 26.8 mM add to M9-agar: 4 ml (final concentration 0.268 µM)
  • Sodium Cyclamate: stocksolution 10 mM 100 µl applied to whatmanpaper in swimming assays, view methods
  • Tryptone: stocksolution 0.5% (w/v) varying amounts used in swimming assays, view methods and notebook of group 1
  • Vanillin: stocksolution 10 mM 100 µl applied to whatmanpaper in swimming assays, view methods

Buffer and Solution

Recipes for stocks of buffers and solutions.

5x M9 salts stock solution

Amount Substance
64 g Na2HPO4 × 7 H2O
15 g KH2PO4
2.5 g NaCl
5.0 g NH4Cl
ad 1000 ml ddH2O
  • Stir until dissolved; sterilize by autoclaving at 121°C.

CaCl2 Buffer for competent cells

Amount Substance Notes
7.351 g CaCl2 × 7 H2O Alternative 0.1 M CaCl2.
15% Glycerol
ad 500 ml ddH2O
  • Stir until dissolved; sterilize by sterile filtration.

50x TAE for agarose gels

Amount Substance
242 g Tris base
Glacial acetic acid
18.6 g EDTA
ddH2O
  • Stir until dissolved. Working concentration is 1x (40 ml of 50x stock ad 2 l ddH2O).

E. coli

Strains

E. coli strain list - List of the working E. coli strains including genotypes, genetic markers, and alleles.

Storage

Storage of E. coli strains - How to keep E. coli strains.

Glycerol Stock

The glycerol stock is prepared as follow:

  • 900 µl over night culture in LB-liquid medium
  • 900 µl 50% sterile glycerol

Glycerol is extremely viscous. Hence, use cut off tips to pipette. Glycerol stocks were vortexed to ensure uniformly mixture and shock frozen in N2 or dry ice. Then, cultures were stored at -80°C. Do NOT forget to fill in the according GMO S1 construction sheet!

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