Team:Bielefeld-Germany/Labjournal/Cultivation

From 2013.igem.org

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<h1>Cultivation</h1>
<h1>Cultivation</h1>
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<br><b><u> Chloramphenicol stock solution</u></b>
<br><b><u> Chloramphenicol stock solution</u></b>
<p>-Solubilize 20 mg mL-1 Chloramphenicol in 100 % Ethanol </p>
<p>-Solubilize 20 mg mL-1 Chloramphenicol in 100 % Ethanol </p>
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<p> - 100 mL 5x M9 salt solution</p>  
<p> - 100 mL 5x M9 salt solution</p>  
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<tr><th><b> 10x M9 salt solution</b></th><th></th><tr>
<tr><th><b> 10x M9 salt solution</b></th><th></th><tr>
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<tr> <td align="center">NaCl</td>      <td align="center">5 g/L</td> </tr>
<tr> <td align="center">NaCl</td>      <td align="center">5 g/L</td> </tr>
<tr> <td align="center">NH4Cl</td>      <td align="center">5 g/L</td> </tr>
<tr> <td align="center">NH4Cl</td>      <td align="center">5 g/L</td> </tr>
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<p> - 20 mL 20 % glucose</p>
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<p> - 20 % glucose
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<tr> <td align="center"><b>20 % Glucose</b></td>  <td align="center">20 % (w/v)glucose</td>  <td align="center">200 g/L</td> </tr>
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<p>  For 500 mL stock solution add 100 g glucose to 440 mL water. Autoclave for 15 at 121° C</p>
<p> - 1 M MgSO4 </p>
<p> - 1 M MgSO4 </p>
<p> - 1 mL biotin (1 mg/mL)</p>
<p> - 1 mL biotin (1 mg/mL)</p>

Revision as of 13:28, 5 September 2013






Cultivation

[Media]

Chloramphenicol stock solution

-Solubilize 20 mg mL-1 Chloramphenicol in 100 % Ethanol

-Store at -20 °C


DNA loading buffer

- 50 % (v/v) glycerol

- 1 mM EDTA

- 0.1 % (w/v) bromphenol blue

- Solve in ddH2O


LB medium

- 10 g Trypton

- 5 g yeast extract

- 10 g NaCl

- 12 g Agar-Agar (for plates)

- Adjust pH to 7.4


M9 medium

For 1 L M9 mineral medium you need 867 mL sterile water (for plates add 16 g Agar-Agar as well). Then add (in the following order:

- 300 µL 1 M CaCl2

- 100 mL 5x M9 salt solution

10x M9 salt solution
Na2HPO4-2H2O 75.2 g/L
KH2PO4 30 g/L
NaCl 5 g/L
NH4Cl 5 g/L

- 20 % glucose

20 % Glucose 20 % (w/v)glucose 200 g/L

For 500 mL stock solution add 100 g glucose to 440 mL water. Autoclave for 15 at 121° C

- 1 M MgSO4

- 1 mL biotin (1 mg/mL)

- 1 mL thiamin (1 mg/mL)

- 10 mL 100x trace elements solution





[Buffers, Solutions]

TAE buffer

For 1 L of 50 x TAE buffer you need:

- 242.48 g Tris

- 41.02 g Sodiumacetate

- 18.612 g EDTA

- Adjust pH to 7.8 with acetic acid

- Solve in dH2O

10 mL of the stock is diluted in 1 L dH2O for the gel electrophoresis (0.5 x TAE buffer)















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