Cetyltrimethylammonium bromide (CTAB) 2% lysis buffer

Here's how to mix a 2% CTAB lysis buffer:

Materials:

  • Cetyltrimethylammonium bromide (CTAB) powder
  • Deionized water (dH2O)
  • 0.5 M Tris-HCl buffer (pH 8.0) (or alternative buffer as per your protocol)
  • 1.4 M NaCl solution
  • 20 mM EDTA solution (optional, for improved RNA isolation)
  • Balance
  • Graduated cylinder
  • Stir plate with stir bar (or equivalent for mixing)
  • Disposable container

Steps:

  1. Calculate amounts:
    • Decide on the final volume of your lysis buffer (e.g., 100mL, 500mL).
    • To make a 2% (w/v) solution, you'll need 2 grams of CTAB per 100 mL of buffer. So, for a 500 mL solution, you'd use 10 grams of CTAB.
  2. Weigh out CTAB:
    • Using a balance, carefully weigh out the required amount of CTAB powder.
  3. Dissolve CTAB in warm water:
    • Add the weighed CTAB to a graduated cylinder containing a small volume of warm dH2O (around 50°C) - this helps CTAB dissolve more easily.
    • Stir the solution using a stir plate with a stir bar until the CTAB dissolves completely. The solution may appear cloudy initially, but it should become clear with proper mixing.
  4. Add remaining components:
    • Once the CTAB is dissolved, add the following solutions to the graduated cylinder in the order mentioned:
      • 0.5 M Tris-HCl buffer (pre-adjusted to pH 8.0)
      • 1.4 M NaCl solution
      • 20 mM EDTA solution (optional)
    • After adding each component, stir the solution thoroughly.
  5. Adjust volume:
    • After adding all the components, bring the final volume of the solution to your desired amount using dH2O.
  6. Store the buffer:
    • Once prepared, aliquot the lysis buffer into smaller volumes and store them at 4°C for short-term use or -20°C for long-term storage.

Safety:

  • CTAB can irritate, so wear gloves and eye protection when handling it.

Additional Tips:

  • Some protocols may call for heating the CTAB solution to higher temperatures (around 65°C) to ensure complete dissolving. However, prolonged heating might affect other components in the buffer. Check your specific protocol for recommendations.
  • Pre-made stock solutions of Tris-HCl buffer, NaCl, and EDTA are often available commercially and can be used for this recipe.
  • Consider your experiment's specific needs. Some protocols may include additional components in the CTAB buffer, like beta-mercaptoethanol for RNA isolation or lysozyme for enhanced cell wall lysis.