TB

Uses

TB buffer is used to make chemically competent cells by the Inoue method Preparing chemically competent cells (Inoue)

Materials

• PIPES 0.5M solution pH 6.7
• Manganese chloride tetrahydrate
• Calcium chloride dihydrate
• Potassium chloride
• Potassium hydroxide 1M solution
• Sterile 0.22 μm bottle top filter
• vacuum source
• clean glassware (no detergent residue -- clean with 70% EtOH)

Preparation

For a 1 liter solution add:

• 250 mM potassium chloride (18.65 g)
• 15 mM calcium chloride (2.2 g)
• 10 mM PIPES (20 ml of an 0.5 M solution)

Adjust pH to 6.7 with KOH.

Add 15 mM manganese chloride (10.88 g)

Sterile filter with a pre-washed 0.22 μm filter and store indefinitely at 4 degrees.