Bromeliaceae
Bromeliad Micropropagation
Bromeliads are tropical plants native to the Americas with wide distribution in rain forests, deserts and coastal areas. Most are epiphytes and terrestrial, diverse and ecologically important, found in microhabitats with strong interactions with fauna.
These protocols focus on Brazilian Atlantic Forest species and demonstrate multiple regenerative pathways including nodule cluster culture, somatic embryogenesis, and temporary immersion systems for mass propagation.
The morphogenetic pathway based on nodule cluster culture is stressed as it appears recurrent in bromeliads, results in high regenerative rates, and is easy to handle. All protocols follow a systematic 5-stage micropropagation process.
General 5-Stage Micropropagation Process
- Weekly application of chelated fertilizer (30-10-10 NPK) and Nitrofoska®
- Apply 10 mL NAA (5 mM) + Vitamin B1 to rosette to enhance offshoot formation
- Spray with benomyl and mineral oil
- Maintain in phytotron: 25±2°C, 16h light (300 μmol/m²/s)
- Isolate axillary buds from young offshoots (1.0-2.5 cm)
- Sterilize: 70% ethanol (2 min) → 1.5% sodium hypochlorite (20 min) → rinse 3×
- Inoculate on paper filter bridges in liquid medium
- Subculture shoots in 300 mL flasks
- Species-specific hormone combinations
- 5-8 clusters per flask in liquid medium
- Sort shoots: <2 cm return to multiplication
- Shoots >2 cm transfer to elongation medium
- Often supplemented with GA₃ (5-10 μM)
- Target size: >3 cm for acclimatization
- Minimum shoot size: 3.0 cm
- Roots optional (prune to 1 cm if present)
- Trays with 128 cells (60 cm³ each)
- Greenhouse with controlled mist, 50% shade
- Survival rates: 85-98%
Base Culture Medium (MS + Morel Vitamins)
| Component | Concentration (mg/L) |
|---|---|
| Macronutrients | |
| NH₄NO₃ | 1,650 |
| KNO₃ | 1,900 |
| CaCl₂·2H₂O | 440 |
| KH₂PO₄ | 170 |
| MgSO₄·7H₂O | 370 |
| Morel Vitamins | |
| Myo-inositol | 100 |
| Thiamine-HCl | 1.0 |
| Pyridoxine-HCl | 1.0 |
| Nicotinic acid | 1.0 |
| Ca pantothenate | 1.0 |
| Glycine | 0.01 |
| Standard Additions | |
| Sucrose | 30 g/L |
| pH | 5.5-5.8 |
| Autoclave | 121°C, 15-16 min |
Note: FeEDTA and micronutrients follow standard MS formulation
Substrate Options for Acclimatization
- Option 1: 2:1:1 (v:v:v) Plantmax® HA : pine bark : carbonized rice coat
- Option 2: 2:2:1 (v:v:v) peat : vermiculite : sand
- Option 3: 1:1 (v:v) carbonized rice coat : Turfa Fértil® mineral supplement
- Maintenance: Weekly spray with ¼MS salts solution
Species-Specific Protocols
Vriesea friburgensis var. paludosa
Nodule Cluster Atlantic Forest
Regeneration from nodule cluster cultures using 2,4-D/Kin induction followed by 2-iP multiplication.
Vriesea reitzii
Endemic Brazil 2 Protocols
Adventitious shoots and nodule cluster regeneration. Red/yellow inflorescences. Threatened species.
Vriesea splendens
Flaming Sword TDZ Protocol
Nodule cluster culture with Thidiazuron multiplication. Brilliant red/orange bracts.
Vriesea fosteriana
TDZ Protocol Encapsulation
Standard protocol plus synthetic seed encapsulation in sodium alginate. Large ornamental species.
Vriesea hieroglyphica
TDZ Protocol Ornamental Foliage
Direct organogenesis with TDZ. Prized for glossy green leaves with dark horizontal bands.
Alcantarea imperialis
Imperial Bromeliad TDZ Protocol
Giant bromeliad (up to 2m). TDZ multiplication protocol. Purple/dark green bracts.
Dyckia distachya
Somatic Embryo Organogenesis
Dual protocols: Picloram-induced somatic embryogenesis and BAP/Kin organogenesis.
Aechmea fasciata
TIS Protocol 30:1 Rate
Temporary immersion system for mass propagation. High multiplication rate (30:1).
Vriesea brusquensis
TIS Protocol Mass Propagation
Temporary immersion system protocol. 3h stationary : 3min immersion cycle.
Contamination Control
- Bacterial: 20 mL/L Kanamycin and/or 100 mg/L Agrimicin® (filter sterilized)
- Fungal: 20 mg/L miconazole nitrate (filter sterilized)
- Prevention: Mother plant spray with benomyl and mineral oil, 1 month in phytotron