IGEM:Virginia 2012/Protocols/Bordet Gengou (BG) Agar Plate Preparation

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This protocol makes the Bordet Gengou Agar Plates, a differential medium used to isolate Bordatella.

Contents

Materials

  • 30g of Bordet Gengou Agar Base
  • 1 L of distilled H2O
  • 13 mL glycerol
  • 2600 mL Fernback Flask
  • Stir Bar
  • 100 mL screw cap bottles
  • 13 mL defibrinated sheep blood
  • Autoclave
  • Autoclave tape
  • 4°C Chamber

Procedure

Making solution

  • Add 30g of Bordet Gengou Agar Base to 1 liter of distilled H2O in a 2600 mL Fernback Flask while stirring.
  • Turn on heat to medium hot.
  • Add 13 mL of glycerol
  • When thoroughly heated and in solution, aliquot 80 mL in to 100 mL screw cap bottles.
  • Leave caps loose enough for air exchange and place autoclave tape between cap and bottle.
  • Autoclave
  • Tighten the caps and store at 4°C when done.

Pour plates

  • Heat autoclaved BG agar bottles in hot water bath until agar is completely liquid.
  • This requires hottest setting on hot plate. Remove bottles to cool on bench top.
  • When comfortable to touch, add 13 mL defibrinated sheep blood and pour into 60 x 15 mm culture dishes, avoiding air bubbles. (If blood is added when BG agar is too hot, it will hemolyze the blood, turning it brown).
  • Note: for antibiotic plates, add 1 mL of 100x solution of antibiotic at time blood is added.
  • When plates have cooled and become solid, turn them upside down.
  • Leave overnight to reduce condensation, then package, date, label, and store at 4°C.

Notes

Please feel free to post comments, questions, or improvements to this protocol. Happy to have your input!


References

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