Tissue Culture Propagation of Begonias

Table of Contents

Begonia is the largest genera of flowering plants with 1,831 species. Members of the group include annuals, perennials, shrubs, and climbers. Some members of the plants also form underground tubers and rhizomes. They occur in subtropical and tropical moist climates in South and Central America, Africa, and Southern Asia.

The leaves of the Begonia plant are often large and variegated. The flowers come in different shapes and colors and are monoecious (male and female flowers of the plant occur separately on the same plant). The fruits are winged capsules containing numerous minute seeds.

Regular watering is required to keep the potting soil slightly moist as the plants are not drought tolerant. For the best bloom and growth, fertilize the plants with a dilute balanced fertilizer throughout the growing season. The problem with the plant is they are susceptible to powdery mildew, botrytis, and stem rot, especially in humid conditions with poor air circulation.

Image credit: https://www.doityourself.com/stry/begonias

Classification

Class: Rosids

Order: Cucurbitales

Family: Begoniaceae

Genus: Begonia

Tissue Culture of Begonia

Begonia plants are popular among cultivators because of their beautiful flowers and, throughout the world, they are used as garden plants, pot plants, hanging basket plants, and decorative indoor plants.

The thin cell layer system (TCL) is an efficient tissue culture technique to obtain a large number of plantlets in a very short time. So, this section presents you with the method for tissue culture of three different explants of Begonia by using the TCL technique.

Material Required

Explants: petiole, stem, and stalks of Begonia tuberous

Culture Media:

• Basal MS media adjusted to pH 5.8 and solidified with 2.5g/L gellan gum.
• Autoclave the prepared media at 121 ℃, 1 atm for 20 minutes.

Culture conditions: Incubate the cultures in growth chambers with environmental conditions adjusted to 23-27 ℃, 75-80% relative humidity, and a photoperiod of 10 hours per day.

Note: Culture the shoots in 100mm x 20mm Petri dishes containing 40ml semi-solid liquid medium.

Procedure

1. Sterilization

• Wash thoroughly the petiole, stem, and floral stalk of Begonia tuberous under tap water for 30 minutes.
• Then, soak all the three parts in detergent for 10 minutes and rinse 6 times with distilled water and once with 70% ethanol.
• Again, wash all three parts with distilled water and then disinfect with a 0.1% HgCl2 aqueous solution for six minutes followed by rinsing six times with distilled water.

2. Thin Cell Layer Cutting

• Segment the Begonia floral stalk into transverse thin cell layers (tTCL), 0.2-1.0 mm discs as shown in the figure below.

Figure: A schematic representation of cutting Begonia floral stalks.

Source: Nhut, D. T., Hai, N. T., & Phan, M. X. (2009).

• Cut the petiole into the transverse thin cell layers (tTCL), 0.2-1.0 mm discs as shown in the figure below.

Figure: A schematic representation of cutting Begonia petiole-leaf.

Source: Nhut, D. T., Hai, N. T., & Phan, M. X. (2009).

• Cut the Begonia main stem transversely (2 mm thick) into several sections and then culture the sections on the MS medium with growth hormones like BA and IAA.

3. Culturing the Explant

Petiole TCLs

1. Culture petiole TCLs on MS media containing 0.2 mg/L TDZ (thidiazuron).
2. Shoot initiation can be observed after 3 weeks of culture.
3. After 8 weeks of shoot initiation, transfer the explants to a fresh MS media without any plant growth regulators for shoot elongation.
4. Then, cut the elongated shoots into small discs and transfer them to a fresh MS medium without any plant growth regulator for further elongation of the shoot.
5. After 2 weeks of elongation, transfer the elongated shoots on MS medium containing 1.0 mg/L BA.

Stem TCLs

1. Culture the thin cell layers of stem on MS media containing 0.2 mg/L BA (6-benzyladenine) and 0.2 mg/L NAA (1-naphthaleneacetic acid).
2. After 4 weeks, shoot generation can be observed.
3. Then, transfer the grown shoots to a fresh MS medium containing 0.2 mg/L BA and 0.2 mg/L NAA.

Floral stalk TCLs

1. Culture the cut discs of floral stalk on MS media containing 0.2 mg/L TDZ (thidiazuron).
2. The generation of shoots can be observed after 3 weeks of culture.

4. Induction of Root Formation

1. When the shoots are grown about 1-1.5 cm in height, with 3-4 leaves, transfer them to a rooting medium.
2. The rooting medium consists of MS medium containing 0.5 mg/L BA, 0.1 mg/L NAA, and 1 g/L activated charcoal after 45 days of culture.
3. After a few weeks, plantlets with developed roots and shoots will be obtained.
4. Then, transfer these plantlets to the greenhouse.

5. Acclimatization

1. After the formation of roots, transfer the plantlets to a sterile soil:sand mixture in 1:1 ratio, pH 5.8, in jam bottles for hardening.
2. Then, after 6 weeks, transfer these plantlets to polybags containing the same potting mixture.
3. Keep the plantlets in the greenhouse for acclimatization at 25 ℃ and 80% humidity.
4. After 6 months, the plants will adapt to the environment well and start flowering.

References

1. Nhut, D. T., Hai, N. T., & Phan, M. X. (2009). A Highly Efficient Protocol for Micropropagation of Begonia tuberous. Methods in Molecular Biology, 15–20. DOI:10.1007/978-1-60327-114-1_2
2. https://www.missouribotanicalgarden.org/PlantFinde...
3. https://extension.umn.edu/flowers/tuberous-begonia...
4. https://en.wikipedia.org/wiki/Begonia