Team:Goettingen/Project/Materials

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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
57.1 ml	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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@@ Line 80: / Line 80: @@
 |242 g || Tris base
 |-
-|57.1 m l| Glacial acetic acid
+|57.1 ml || Glacial acetic acid
 |-
 |18.6 g || EDTA