Aloe vera propagation is typically done through ramets or bud insertion, which are time-consuming and seasonally restricted. However, using tissue culture techniques offers a more efficient and year-round solution. This method involves inducing callus and shoot formation from explants, followed by subculturing to produce a large number of genetically identical plants. It's especially beneficial for introducing new varieties, as it ensures consistent quality and rapid production.
**Materials and Methods**
1. **Plant Species**: Aloe vera
2. **Explant Source**: Young stem segments
3. **Culture Conditions**:
- Basic medium: MS (Murashige and Skoog)
- Hormones:
- 0.5–3 mg/L 1BA (cytokinin) + 0.01–0.5 mg/L NAA (naphthalene acetic acid)
- 0.5–1 mg/L 2BA + 0.01–0.1 mg/L NAA
- 0.2–1 mg/L GA3 (gibberellin)
- 0.2–0.4 mg/L IBA (indole-3-butyric acid) + 1 mg/L CCC (chlormequat chloride)
- Temperature: 23–27°C
- Light: 1500–2000 lux, 10 hours per day
4. **Experimental Procedure**:
- Collect young stems and disinfect them before cutting into 0.5–1 cm segments on a sterile bench.
- Inoculate the segments into medium 1 for callus induction. After 2–3 weeks, callus and small shoots begin to form.
- Transfer to medium 2 after one week, where clustered shoots develop rapidly.
- When shoots reach 1.5–2 cm in height, they are separated and transferred to medium 3 for root development.
- After three weeks, fully developed plantlets with 3–5 cm height and 2 cm roots are ready for transplanting.
- For mass propagation, callus can be subcultured on medium 1 to expand the population.
**Clone Establishment and Storage**
Aloe callus can be stored at 5°C for up to five months without losing viability. Before use, stored callus should be cultured in the lab for three days to check for contamination. If no contamination is found, the callus can be expanded. The medium remains viable for long-term storage if kept fresh.
**Transplanting and Management of Test Tube Seedlings**
Test tube seedlings have weak adaptability to external conditions and require careful acclimatization. Before transplanting, the lid should be opened gradually, and the seedlings left in the growth room for 2–3 days. When removing them, avoid damaging the roots, and gently wash the roots in room temperature water to prevent rotting.
Use a mix of garden soil and vermiculite or pure vermiculite as the substrate. After planting, cover the seedlings with a thin plastic film, mist daily for 30 minutes, and transplant after four weeks. This process ensures better survival and faster establishment in the field.
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