The Latest Propagation Techniques and Methods Tutorial for Anoectochilus

The traditional propagation methods for Anoectochilus are division, cuttings, and seed sowing. Among these, because seeds are difficult to harvest and the germination rate of sowing is low, division and cuttings were mostly used for propagation in the past. However, because this method is easily affected by the environment, its survival rate is about 30~70%. Large-scale cultivation generally uses tissue culture for propagation. Let us take a look together below!

Division Propagation of Anoectochilus

Division propagation of Anoectochilus can be carried out in both spring and autumn, generally once every three years. All plants that grow robustly and have dense pseudobulbs can be divided. After division, each clump must retain at least 5 pseudobulbs connected together. Before division, watering should be reduced to make the potting soil relatively dry. When potting after division, first cover the drainage hole at the bottom of the pot with broken tile pieces, then spread coarse gravel to occupy 1/5 to 1/4 of the pot depth, then add coarse-grained soil and a small amount of fine soil, and then plant with sandy loam rich in humus. The planting depth should be such that the pseudobulbs are just buried in the soil, leave a 2 cm rim at the edge of the pot, spread Selaginella uncinata or fine gravel on top, and finally water thoroughly, place in a shady area for 10-15 days, keep the soil moist, gradually reduce watering, and carry out normal care.

Seed Propagation of Anoectochilus

The seeds of Anoectochilus are extremely fine, and the seed contains only one incompletely developed embryo, so its germination capacity is very low. In addition, the seed coat does not easily absorb water, so sowing by conventional methods cannot germinate. Therefore, orchid mycorrhizal fungi or artificial culture medium are needed to supply nutrients for germination. For sowing, it is best to select fruit that has not yet split open. After sterilizing the surface with 75% alcohol, take out the seeds, soak them in 10% sodium hypochlorite for 5-10 minutes, take them out and rinse them 3 times with sterile water, then sow them in a culture bottle containing medium, and then place them in a dark culture room with the temperature kept at about 25C. After they begin to sprout, move them into the light and they can form protocorms. From sowing to transplanting, Anoectochilus takes six months to one year. Tissue culture has already been successful, and this method can be used for propagation where conditions permit.

Tissue Culture of Anoectochilus

1. Prepare the culture medium: Prepare the culture medium according to the specific formula requirements. Package the prepared medium well and place it in an autoclave for sterilization. The sterilized medium can either be used directly or stored in a sterile inoculation room for later use, but the storage time should not exceed two weeks.

2. Cut explants and sterilize for inoculation: Under sterile conditions, cut explants of appropriate length from excellent mother plants (stem tips, stem segments, leaves, buds, roots), inoculate them into the culture medium, and place them in the culture room for induction.

3. Clustered shoot induction and differentiation induction: Under conditions of 20-28°C in the culture room and 12-16 hours of illumination from fluorescent lamps with a light intensity of 3000-4000 Lux, and at the same time under the action of high-concentration cytokinins and low-concentration auxins in the culture medium, the explants are induced to differentiate clustered shoots.

4. Subculture proliferation:
After the clustered shoots grow in the culture medium for a period of time (usually 20-30 days), as the nutrients in the medium are consumed, they must be transferred to fresh medium for culture, that is, subculture. In order to expand the propagation speed and propagation coefficient, subculture transfer should be carried out. By changing the culture conditions, such as culturing for a few days at a lower temperature and then transferring to another higher temperature for a few days, or by liquid shaking culture for a certain period of time, the proliferation speed can be significantly increased.

5. Rooting induction: When the shoots grow to a certain height, transfer them to rooting medium with a higher auxin concentration to continue culturing. First carry out dark culture on the young shoots for one week, then transfer them to normal light, or shorten the light period, which can advance rooting and increase the rooting rate and rooting speed.

6. Transplanting and hardening of seedlings: Move rooted seedlings from the culture room to outdoor soil and cultivate them into qualified afforestation
seedlings.

① Seedling grading: According to the quality indicators of seedlings, grade the seedlings. Grade I and Grade II seedlings should be transplanted in a timely manner, while Grade III seedlings should continue to be cultured.

② Hardening: Usually use the method of increasing light for about one week of hardening, or add retardants to the culture medium, and 1-2 days before transplanting move the seedlings together with the culture bottles from the culture room to the greenhouse, and open the bottle stoppers.

③ Transplanting: When the root system grows to about 1 cm, carefully take the seedlings out of the culture bottles, wash off the culture medium with clean water, and be sure not to damage the plants. First soak the seedlings in a relatively dilute sodium sulfate solution for a few minutes before transplanting, spray water thoroughly, and cover with plastic film to prevent wind and preserve moisture.

④ Management after transplanting: In the early stage after transplanting, provide appropriate shading to prevent direct sunlight, then gradually increase the light duration and intensity, keep the temperature at 20-30°C, water appropriately to preserve moisture, control soil moisture, and gradually implement full light. After hardening for a period of time, when the seedlings have fully adapted to the natural environmental conditions, they can be transplanted into lightweight substrate mesh bags, and when the seedlings meet the standards, they can be released from the nursery.