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Tissue Culture Media Quick Guide

Last updated: April 29, 2025 Plant Science & Propagation

Contents

Common Media Types

These commercially available formulations are most commonly used for cannabis tissue culture:

Media TypeBest ForSupplier OptionsNotes
MS (Murashige & Skoog)General cannabis culturePhytoTech, Sigma, Caisson LabsMost widely used, suitable for most cannabis varieties
½ MSRoot initiationSame as MSUse half-strength MS for rooting
Gamborg B5Callus and suspension culturesPhytoTech, Sigma, Caisson LabsLower nitrogen content, less ammonium
DKWWoody plants/cannabisPhytoTechSometimes preferred for mature tissue
WPM (Woody Plant Medium)More difficult genotypesPhytoTech, SigmaAlternative for strains sensitive to MS

Using Pre-Mixed Media Sachets

Pre-mixed media sachets are the simplest and most reliable approach for most growers.

What You’ll Need

  • Pre-mixed media powder (MS, B5, etc.)
  • Sucrose (table sugar works fine)
  • Gelling agent (agar or gellan gum)
  • pH meter or pH test strips
  • pH adjustment solutions (0.1M NaOH and 0.1M HCl)
  • Distilled or reverse osmosis water
  • Plant growth regulators (if needed)

Standard Formulation (1 Liter)

ComponentAmountNotes
Pre-mixed media powder1 packet (or as directed)Follow supplier’s instructions
Sucrose20-30g2-3% concentration (3% standard)
Agar6-8gFor solid medium
OR Gellan gum (Gelrite/Phytagel)2-2.5gClearer medium than agar
Final pH5.8-5.9Adjust before adding gelling agent

Cannabis-Specific Modifications

Simple additions to pre-mixed media that can improve cannabis culture success:

AdditionAmount Per LiterBenefit
Activated charcoal0.5-1.0gReduces browning, absorbs toxins
Ascorbic acid50-100mgAntioxidant, prevents browning
Cysteine10-20mgAntioxidant, prevents browning
PVP (polyvinylpyrrolidone)250-500mgBinds phenolics, reduces browning
Additional calcium (CaClâ‚‚)10-20% extraImproves structural integrity
Silicon (K₂SiO₃)50-100mgStrengthens cell walls

Tip: For cannabis, slightly reducing nitrogen (using ½ strength MS) often produces better results with lower callusing and better morphogenesis.

Simple Media Preparation Protocol

Quick Method (1 Liter)

  1. Gather Supplies

    • 1L heat-resistant container/flask
    • Pre-mixed media sachet (MS, B5, etc.)
    • Digital scale
    • pH meter or test strips
    • Sucrose (regular table sugar works)
    • Agar or gellan gum
    • Distilled or RO water

  2. Preparation Steps

    1. Add ~800ml distilled water to container
    2. Add pre-mixed media powder and stir until dissolved
    3. Add 20-30g sucrose (2-3%) and stir until dissolved
    4. Add any plant growth regulators or supplements (if needed)
    5. Check and adjust pH to 5.8-5.9 using pH up/down solutions
    6. Add water to make ~950ml total volume
    7. Add 6-8g agar or 2-2.5g gellan gum while stirring
    8. Add water to reach 1L total

  3. Sterilization

    • Cover container with aluminum foil
    • Autoclave at 121°C (15 psi) for 15-20 minutes
    • If using a pressure cooker: 15 psi for 20 minutes

  4. Pouring

    • Cool to about 60°C (hot but touchable)
    • Pour 25-30ml per culture vessel in a clean environment
    • Allow to solidify with lids slightly ajar to prevent condensation
    • Once cool, close vessels tightly

Common Growth Regulators for Cannabis

StageHormonesConcentrationPurpose
InitiationBAP/BA0.5-1.0 mg/LShoot induction
MultiplicationBAP/BA0.2-0.5 mg/LShoot proliferation
RootingIBA or NAA0.1-0.5 mg/LRoot induction
Callus2,4-D + BAP0.5-2.0 mg/L eachCallus induction

Troubleshooting

ProblemLikely CauseQuick Fix
Media not solidifyingpH too high or overheatedLower pH slightly, use fresh agar
Excessive condensationPoured too hotLet vessels cool with lids ajar
Cultures browningPhenolic oxidationAdd activated charcoal or antioxidants
ContaminationPoor sterile techniqueUse PPMâ„¢ (1-2ml/L) as antimicrobial
Hyperhydricity (glassy tissue)Too humid, cytokinin too highReduce humidity, lower BAP concentration