Plant Tissue Culture : Equipment, Steps and Techniques

Published on 16-Aug-2025

Equipment and Materials Required for Tissue Culture

Equipment Required

  • Washing bucket

  • Balance

  • pH meter

  • Autoclave

  • Microscope

  • Microtome

  • Refrigerator

  • Laminar air flow cabinet

  • Centrifuge machine

  • Microwave oven

  • Shaker

  • Burner

Glassware

  • Test tubes

  • Conical flasks

  • Cylindrical flasks

  • Beakers

  • Petridis

  • Measuring cups

  • Pipettes

  • Seeded glass rods

  • Bottles

Chemicals

  • Various inorganic salts

  • Growth hormones

  • Vitamins

  • Amino acids

  • Charcoal

  • Yeast

  • Alcohol

  • Spirits

  • Carbohydrates

  • Etc., for preparing the culture medium

Other Materials

  • Scissors

  • Tweezers

  • Surgical bread scissors

  • Spatula

  • Filter paper

  • Aluminum foil

  • Plastic containers

Sterile Environment

Tissue culture requires a completely sterile environment for proper processing.

  • Culture medium containers and glassware: Sterilized in a hot air oven at 160–180°C for 1–2 hours.

  • Instruments (forceps, needles, scalpels, etc.): Sterilized by heating with a burner and immersing in 95% alcohol.

Types of Tissue Culture

  1. Axillary bud culture

  2. Meristem culture

  3. Micropropagation

  4. Seedling production through callus culture

  5. Somatic embryogenesis

  6. Haploid through pollination culture

  7. Plant production

  8. Protoplast culture

Steps in the Tissue Culture Procedure

Step 1: Selection of Mother Plant or Explant

  • Explant: Part of a plant separated for tissue culture.

  • Top bud, lateral bud, or leaf tip is commonly used.

  • The mother plant must be disease-free and of excellent quality.

Step 2: Preparation of Culture Medium

  • Combine all chemical elements required for plant nutrition and growth.

  • Include macroelements, microelements, vitamins, sucrose (2–4%), phytohormones.

  • Add coagulating material (agar) to thicken the medium.

  • Basal medium: Rich in basic elements, pH 5.5–5.8.

Step 3: Sterilization

  • Medium and explants must be sterilized.

  • Medium poured into flasks or test tubes, closed with sterile cotton, sterilized in an autoclave at:

    • Temperature: 121°C

    • Pressure: 15 lb

    • Time: 20 minutes

  • In-vitro culture: Artificial cultivation of explants under sterile conditions.

Step 4: Placement of Explants in Medium

  • Explants sterilized with alcohol and placed in sterile glassware containing culture medium.

Step 5: Callus Formation and Multiplication

  • Place explants in a controlled environment:

    • Light: 3,000–5,000 lux

    • Temperature: 17–20°C

    • Humidity: 70–75%

  • Tissue divides to form a shapeless mass called callus.

  • Numerous buds form from the callus.

Step 6: Transplantation and Seedling Production in Root Production Medium

  • Buds carefully cut and placed in root production medium.

  • Each bud develops into a full-fledged plant.

Step 7: Transplanting Seedlings into Tubs

  • Well-formed seedlings removed from culture pot and transferred carefully to tubs.

Step 8: Seedling Storage

  • Seedlings kept in humid conditions for acclimatization.

  • Once strong, transplanted into natural soil environment.

Applications of Tissue Culture Technology

Plant Breeding and Advanced Innovation

  • Micropropagation: Producing many seedlings from small tissue for plants that cannot produce seeds (e.g., Thuja, Sea Banana).

  • Seedlings produced are genetically identical to parent plants.

  • Tissue culture allows large-scale production of quality plants for flowers, fruits, and grains.

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