5-triiodobenzoic acid

5-Triiodobenzoic Acid in Plant Tissue Culture

Safety Note: Always consult the SDS for 5-triiodobenzoic acid and follow institutional safety procedures; treat unknowns conservatively. 5-Triiodobenzoic acid is not known to be mutagenic or carcinogenic, but it is a chemical compound and should be handled with appropriate safety precautions, including wearing gloves and eye protection.

Overview and Identity

5-Triiodobenzoic acid (TIBA) is a synthetic auxin transport inhibitor used in plant tissue culture to manipulate plant development. It’s not a nutrient, vitamin, or buffer, but rather functions as a plant growth regulator (PGR) by interfering with the polar transport of endogenous auxins.

Common Names, Synonyms, and Abbreviations

Common name: 5-Triiodobenzoic acid
Synonyms: 2,4,6-Triiodo-benzoic acid; Triiodobenzoic acid
Abbreviations: TIBA

Chemical Identity

Formula: C₇H₃I₃O₂
Relevant forms: Typically available as a solid powder, usually in tissue-culture-grade purity. Hydrate states vary depending on supplier and storage conditions.

Functional Role(s) in Plant Tissue Culture

TIBA’s primary function is as a plant growth regulator (PGR), specifically an auxin transport inhibitor. It doesn’t act as a macronutrient, micronutrient, vitamin, buffer, chelator, gelling agent, sterilant, solvent, or mutagen.

Mechanism and Rationale in vitro

TIBA inhibits the polar auxin transport system within plant tissues, blocking the basipetal movement of auxin. This leads to an accumulation of auxin in the apical regions and a reduction in auxin concentration in the basal regions. The effect is dependent on concentration and the plant species. By altering the distribution of auxin, TIBA influences various developmental processes.

Stage-Specific Relevance

• Shoot proliferation: TIBA can promote shoot proliferation by preventing the apical dominance normally enforced by auxin accumulation. This can lead to increased branching and the development of multiple shoots from a single explant.
• Rooting: While primarily an auxin transport inhibitor, its effects on rooting are complex and species-dependent. Low concentrations may sometimes promote rooting by altering the auxin gradient, while higher concentrations frequently inhibit root formation.
• Somatic embryogenesis: TIBA’s use in somatic embryogenesis is not standard practice. However, indirect influence on auxin distribution might affect embryo development in some species.
• Callus induction: TIBA is frequently used in conjunction with auxins (2,4-D, NAA) to modify callus development and morphology.
• Protoplasts: No established use in protoplast culture.
• Contamination control: TIBA has no direct antimicrobial action.

Interactions or Compatibility/Antagonism with Other Agents

TIBA acts primarily by interacting with the plant’s endogenous auxin system. Therefore, its effects are highly dependent on the auxin/cytokinin balance in the medium. The presence and concentrations of other growth regulators will influence the response to TIBA. Compatibility with other medium components is generally good, provided appropriate solubility is ensured. It may affect gellan gum gel strength at high concentrations.

Preparation and Stock Solutions

Solubility: TIBA is sparingly soluble in water. It is more readily dissolved in alkaline solution (e.g. 0.1N NaOH). Ethanol or DMSO can be used as solvents, but their use requires careful consideration of plant tolerance.

Typical stock concentrations: Stock solutions are typically prepared at 1000mg/L (1g/L) or higher.

Preparation: Accurately weigh the required amount of TIBA. Add a small volume of the chosen solvent, or 0.1N NaOH, to dissolve the powder. Add remaining solvent to reach final volume. Adjust pH if needed, and filter sterilize (0.22 µm) before adding to the cooled, autoclaved medium. TIBA, itself, is readily autoclavable, unlike some other plant growth regulators.

Example stock recipe (1g/L):

1. Weigh 1g of tissue-culture-grade TIBA.
2. Add 50 mL of 0.1N NaOH and stir until dissolved.
3. Make up to 1000 mL with sterile distilled water.
4. Filter sterilize (0.22 µm).
5. Store in amber glass bottle at 4°C.

Working Concentrations and Usage in Media

Common working concentration range: 0.1-10 mg/L, depending on the species and the specific application. It’s crucial to conduct dose-response experiments to determine the optimal concentration for a particular plant species and explant. Add TIBA to the cooled, autoclaved medium after filter sterilization.

Storage and Stability

Storage conditions: Store stock solutions in amber glass bottles at 4°C, protected from light and moisture.

Shelf-life: The stability of TIBA stock solutions is variable; conduct regular quality checks and consider empirically-determined expiry periods.

Container type: Amber glass bottles are preferred to minimize photodegradation.

Quality, Sourcing, and Compatibility

Recommended grade: Always use tissue culture-tested grade TIBA. Confirm the purity and perform necessary tests before use to ensure reliability. Lot-to-lot variability can occur; thorough documentation is crucial.

Safety and Precautions

Hazards: Iodinated compounds should be handled with caution. While not known to be highly toxic, skin and eye contact should be avoided. Use a fume hood when preparing large volumes.

PPE: Wear gloves, eye protection, and a lab coat. Follow institutional safety protocols.

Troubleshooting and Optimization

Common issues: Precipitation, inconsistent growth responses, and undesired effects on morphogenesis are potential problems. Troubleshooting revolves around optimizing TIBA concentration, pH, and interactions with other media components.

Example Protocols and Parameters

• Shoot proliferation in Arabidopsis thaliana: 0.5–1mg/L TIBA in shoot proliferation media containing BAP (0.5–1mg/L). Monitor response and adjust accordingly.
• Rooting in Nicotiana tabacum: Lower concentrations (0.1-0.5mg/L) of TIBA might enhance rooting when used in media with appropriate auxins (IBA or NAA). Ranges are species- and explant-dependent; optimize empirically.
• Callus induction in Solanum lycopersicum: 0.5-2 mg/L TIBA in combination with 2,4-D (1-2mg/L), optimize auxin:TIBA ratios empirically.

Documentation and Labeling

Thoroughly document all aspects: chemical form, lot number, preparation date, stock concentration, solvent, pH, storage conditions, expiry date, media batch number, and treatment matrices.

Key Takeaways

• TIBA is an auxin transport inhibitor used to modulate plant development in vitro.
• Its effects are highly species and concentration-dependent; optimization through dose–response experiments is crucial.
• Stock solutions should be prepared using appropriate solvents, filter sterilized, and stored under appropriate temperature and light conditions.
• Always prioritize safety; follow all relevant safety protocols and consult the SDS.