TAP (Tris-Acetate-Phosphate)

TAP salts solution (×40)

per 1 L · usage rate 25 mL per 1 L of medium · storage at 4 °C, no more than 1 month

TAP phosphate solution

per 100 mL · usage rate 0.375 mL per 1 L of medium · storage at 4 °C, 6 months

Hutner's trace elements solution

per 100 mL of solution · usage rate 1 mL per 1 L of medium · storage at 4 °C

Dissolve Tris in 900 mL of water → add 25 mL of the TAP salts solution → 0.375 mL of the phosphate solution → 1 mL of Hutner's solution → add acetic acid dropwise to pH 7.0 (usually about 1.0 mL) → make up to 1000 mL → autoclave.

• The acetic acid has a dual role: it is the source of organic carbon for mixotrophic growth and the agent for adjusting the pH. Hence it is added at the very end, under pH-meter control.
• The salts solution and the phosphate solution are stored separately: in a single bottle calcium and phosphate immediately form a precipitate.
• Variants: TP — the same composition without acetic acid (photoautotrophic growth only); TAP-N — without NH₄Cl (induction of lipid and starch accumulation).
• Because of the acetate, TAP is exceptionally prone to contamination by bacteria and fungi — strict aseptic technique and work in a laminar flow cabinet are mandatory.
• Molar equivalent of the buffer: Tris — 20 mM.
• This is a macronutrient salt concentrate with a high usage rate (25 mL/L), which is why it is prepared per litre rather than per 100 mL: one litre is enough for 40 L of medium. Turbidity indicates bacterial growth; such a solution is discarded.
• The solution is very concentrated; dissolve the salts with warming to 40 °C and constant stirring. Store it separately from the TAP salts solution — combining it with calcium produces a calcium phosphate precipitate.
• A special procedure is required: all the salts except EDTA and FeSO₄ are dissolved in 55 mL of hot water; Na₂EDTA separately in 25 mL; FeSO₄ in 5 mL. The solutions are combined while hot, the pH is brought to 6.7 with hot 20 % KOH, the volume is made up to 100 mL and the flask is left loosely stoppered for 1–2 weeks at room temperature. During this time the colour changes from green to purple and a rusty precipitate forms — filter it off. The solution remains usable for as long as it stays purple-violet.