Loading

Vriesea fosteriana
Large Ornamental

TDZ ProtocolPBZ: 4 μMEncapsulation Available

Vriesea fosteriana is a large bromeliad species spreading from Brazil to Mexico. It shows broad, mid-green leaves with a reddish-brown band on both sides. The foliage can grow up to 1-m long forming a dense rosette. The reference workflow follows the LFDGV/CCA/UFSC benchmark described in Jain & Ochatt (2010).

From the center a conspicuous flower stalk reaches upwards to 1.5 m, developing a yellow flower from each bract. This species is highly valued in ornamental horticulture.

Two protocols are provided: standard TDZ multiplication for mass propagation, and an innovative encapsulation protocol for synthetic seed production and long-term storage.

Basal Medium Reference (MS + Morel)

Both protocols use Murashige & Skoog salts plus Morel vitamins as the basal medium.

Component Stock Volume Final in 1 L
NH₄NO₃ 82.5 g/L 20 mL 1,650 mg/L
KNO₃ 95.0 g/L 20 mL 1,900 mg/L
CaCl₂·2H₂O 88.0 g/L 5 mL 440 mg/L
KH₂PO₄ 34 g/L 5 mL 170 mg/L
MgSO₄·7H₂O 74 g/L 5 mL 370 mg/L
Fe-EDTA solution 37.3 & 27.8 g/L 10 mL Fe chelate pair
Micronutrient mix 1.24–0.0055 g/L 5 mL Boron, iodine, molybdenum, cobalt
Morel vitamins 1–100 mg/L full strength Thiamine, pyridoxine, nicotinic acid, Ca pantothenate, myo-inositol, glycine
Sucrose - - 30 g/L

Standard Protocol Timeline (PDF Benchmark)

Stage 1 · Weeks 0–28

Induction

NAA 2 μM + BAP 4 μM, 3× subculture at 6-week intervals

Stage 2 · Variable

Multiplication

TDZ 0.1 μM, 14:1 multiplication rate

Stage 3 · Weeks 28–38

Elongation

PGR-free MS medium

Stage 4 · Weeks 38–47

Acclimatization

2:1:1 substrate, 98% survival

Standard Protocol: Direct Organogenesis with TDZ

Stage 1: Induction (10 weeks + 18 weeks subculture)

  1. Step 1 Inoculate buds over filter paper bridges in test tubes (25 mm × 150 mm).
  2. Step 2 Use 15 mL liquid medium: MS + Morel vitamins + Sucrose 30 g/L + NAA (2 μM) + BAP (4 μM)
  3. Step 3 After , nodule cluster cultures arise.
  4. Step 4 Subculture 3 times, every 6 weeks, in same medium (total 18 more weeks).

Stage 2: Multiplication with TDZ

  1. Step 1 Subculture 5-8 microshoot clusters per 300 mL flask.
  2. Step 2 Use 15 mL liquid medium per flask.
  3. Step 3 Medium: MS + Morel vitamins + Sucrose 30 g/L + TDZ (0.1 μM)
  4. Step 4 Multiplication rate: 14:1

Stage 3: Elongation (10 weeks)

  1. Step 1 Subculture 8-10 microshoot clusters per 300 mL flask.
  2. Step 2 Use 15 mL liquid medium per flask.
  3. Step 3 Medium: MS + Morel vitamins + Sucrose 30 g/L, free of PGR

Stage 4: Acclimatization (9 weeks)

  1. Step 1 Transfer shoots ≥3.0 cm to trays of 128 cells (60 cm³ each).
  2. Step 2 Substrate: 2:1:1 (v:v:v) Plantmax® HA : pine bark : carbonized rice coat
  3. Step 3 Nebulization tunnel, .
  4. Step 4 Survival rate: 98%

Encapsulation Protocol: Synthetic Seed Production

Preparation: Standard Multiplication (24 weeks)

  1. Step 1 Follow standard induction protocol (see above).
  2. Step 2 After 3-4 subcultures (5-7 weeks each), achieve multiple shoot proliferation.

Induction of Encapsulated Units (8 weeks)

  1. Step 1 Subculture 5-8 clusters in 300 mL flasks with 20 mL medium.
  2. Step 2 Medium: MS + Morel vitamins + Sucrose 30 g/L + NAA (2 μM) + BAP (4 μM) + Paclobutrazol (4.0 μM)
  3. Step 3 After , multiplication rate reaches 13:1.
  4. Step 4 Select shoots 0.5-1.0 cm long for encapsulation.

Encapsulation Process

  1. Step 1 Prepare alginate matrix: 1% sodium alginate in ½ strength MS salts + GA₃ (5 μM)
  2. Step 2 Immerse individual shoots (0.5-1.0 cm) in alginate solution.
  3. Step 3 Complexation: Drop alginate-coated shoots into 50 mM CaCl₂ solution.
  4. Step 4 Maintain in CaCl₂ for to form gel beads.
  5. Step 5 Wash encapsulated units thoroughly in tap water.
  6. Step 6 Storage: Store capsules at 5°C for .

Decomplexation & Sowing

  1. Step 1 Treat capsules with 100 mM KNO₃ solution for before sowing.
  2. Step 2 Transfer synthetic seeds to trays of 220 cells (13 cm³ each).
  3. Step 3 Substrate: Vermiculite

Ex Vitro Conversion in Phytotron

  1. Step 1 Place trays inside plastic boxes covered with glass to allow light entry and reduce water exchange.
  2. Step 2 Conditions: 25°C ± 2°C, 16h light (400 μmol/m²/s)
  3. Step 3 Periodically wet germinating capsules with ¼ MS salts solution.
  4. Step 4 After , transfer trays to greenhouse.

Final Acclimatization (8 weeks)

  1. Step 1 Transfer plantlets to trays of 128 cells (60 cm³ each).
  2. Step 2 Substrate: 1:1 (v:v) carbonized rice coat + Turfa Fértil® (N:4%, P₂O₅:14%, K₂O:8%)
  3. Step 3 Nebulization tunnel with intermittent mist.
  4. Step 4 Duration:
  5. Step 5 Survival rate: 85%
  6. Step 6 Transfer to pots and maintain in greenhouse for .

Encapsulation Timeline (PDF Benchmark)

Prep · Weeks 0–24

Standard Multiplication

3-4 subcultures at 5-7 week intervals

Stage 1 · Weeks 24–32

PBZ Induction

NAA 2 μM + BAP 4 μM + PBZ 4 μM, 13:1 rate

Stage 2 · Day 1

Encapsulation

1% alginate + ½ MS + GA₃ 5 μM, 10 min CaCl₂

Stage 3 · Weeks 32–36

Cold Storage

5°C for 4 weeks

Stage 4 · Weeks 36–41

Ex Vitro Conversion

Vermiculite in phytotron, ¼ MS foliar

Stage 5 · Weeks 41–49

Final Acclimatization

85% survival, then 10 more weeks in greenhouse

Encapsulation Matrix

Critical steps for synthetic seed production from the PDF benchmark.

Step Composition
Alginate Matrix 1% sodium alginate in ½ MS + GA₃ 5 μM
Complexation 50 mM CaCl₂ for 10 minutes
Decomplexation 100 mM KNO₃ for 20 minutes before sowing
Sowing Substrate Vermiculite in 220-cell trays (13 cm³ each)

Protocol Comparison

Choose standard protocol for maximum survival or encapsulation for storage and transport.

Aspect Standard TDZ Encapsulation
Multiplication Rate 14:1 13:1
Survival Rate 98% 85%
Storage Capability Limited 4 weeks at 5°C
Acclimatization In vitro → ex vitro Direct ex vitro
Key Advantage Higher survival Storage & transport
Best For Mass propagation Germplasm conservation

Key Success Factors

  • TDZ efficiency: Very low concentration (0.1 μM) reduces costs significantly vs BAP/NAA
  • PBZ treatment: Paclobutrazol produces compact shoots ideal for encapsulation
  • Shoot size for encapsulation: 0.5-1.0 cm optimal for alginate bead formation
  • Complexation time: Exactly 10 minutes in CaCl₂ ensures proper gel formation
  • Cold storage: 4 weeks at 5°C extends viability and synchronizes germination
  • Decomplexation: KNO₃ treatment essential before sowing to soften alginate matrix
  • Ex vitro germination: Vermiculite + phytotron conditions allow direct conversion without traditional tissue culture
Guerra & Dal Vesco (2010) in Jain & Ochatt (2010), Methods in Molecular Biology, vol. 589