Most higher plants have the capacity to reproduce sexually leading to seed formation resulting in production of genetically different offspring. Plants also have the capacity to reproduce vegetatively resulting in plants having the same genetic composition (clones). Horticulturally, vegetative propagation allows for the efficient cloning of plants while retaining valuable traits. The diverse methods by which plants naturally reproduce vegetatively are fascinating! In this column we will explore interesting propagation methods and examples
So, let’s start our journey by describing the most technologically advanced propagation method called micropropagation. This method is defined as the rapid vegetative multiplication of plants, from shoot-tips or other tissues, cultured in vitro (latin for “in glass” in culture vessels) under sterile conditions on a nutrient medium under controlled conditions of light and temperature. Micropropagation is often referred to as in vitro propagation or plant tissue culture. The benefits of micropropagation include rapid propagation of disease-eradicated plants. Since the early 1960s, micropropagation has become a well-established practice for commercial large-scale production of many horticultural plants, especially ornamental species. Today, there are approximately 70 commercial laboratories in the United States. Some of the larger laboratories are located here in Florida. Most potted house plants you buy at Home Depot, Lowes, or your local garden shop were probably produced in a Florida micropropagation lab! While ornamental plant segment accounts for the highest production, medicinal plants and food crops such potato, bananas, strawberries and blueberries are produced using this technology. In 2020, the global plant market was estimated to be $382 million (Srivas, A. and O. Sumant, 2021).
Plant production through micropropagation is usually achieved through five sequential stages [Stage 0 – IV] (Fig. 1). Depending on the plant and media, new plants may be produced by branching of cultured shoot-tips, formation of adventitious shoots from single cells or even formation of vegetative embryos. Propagation from shoot-tips is the most commonly used method. These stages have been widely adopted by commercial micropropagation industry because they describe procedural steps in the micropropagation process and also represent points at which the culture environment and medium needs to be changed. Commercial laboratories are designed to facilitate efficient completion of these stages. The sequential micropropagation stages are shown in Fig. 1.
In Stage 0, healthy disease-free donor plants are selected (Fig. 2a). To initiate Stage I, small shoot-tips or nodes (called explants) are harvested from the donor plant (Fig. 2b), surface sterilized in a dilute bleach solution (Fig. 2c), rinsed in sterile water and then inoculated onto a sterile medium containing mineral salts, vitamins, sucrose and a mixture of a cytokinin and auxin [plant growth regulators] (Fig. 2d). Most media are solidified with a gelling agent such as agar. Stage I cultures are maintained for about 4 weeks and then indexed for bacterial/fungal contamination (Fig. 2f-g). Contaminated cultures are discarded while those that pass are transferred to a Stage II multiplication medium containing higher cytokinin levels to promote rapid shoot multiplication (Fig. 3a). Stage II shoots cultures can be repeatedly subdivided and continually transferred onto fresh Stage II medium to increase culture numbers. Consider this, if you obtain 10 new shoots from a single original shoot every four weeks, after six months about 1,000,00 plants are produced! Establishment of indexed slowly multiplying cultures maintained long-term as mother block cultures (Fig. 1) provides a secure backup to overcome potential culture loss.
For production, Stage II shoot clusters can be separated into individual shoots called microcuttings (Fig. 3b). Depending on the plant, Stage III, the pretransplant/rooting stage, can be skipped with microcuttings being rooted directly in potting soil under greenhouse conditions. This saves about 65% of the production course, If not, the microcuttings are transferred to a Stage III medium which may contain an auxin to induce rooting (Fig. 3c,d). The rooted microcuttings (Fig. 3d) are removed from the medium, rinsed to remove medium clinging to the roots and then transferred into potting medium and placed under intermittent mist and partial shade to acclimatize plants to growing in greenhouse conditions of lower humidity and higher light levels.
In the US, micropropagation technology is used to facilitate plant, production breeding, and conservation programs at many botanical gardens including Denver Botanical Gardens, Longwood Gardens, and Atlantic Botanical Garden.
Fig. 2. Strawberry micropropagation. a. strawberry donor plant runner, b. runner node with isolated shoot bud (right), c. surface sterilization with dilute bleach, d Stage I inoculation, e. established Stage I culture, f-g. indexing Stage I culture for bacteria/fungal contamination.
Fig. 3. Strawberry micropropagation. a. Stage II shoot cluster after 4 weeks, b. unrooted Stage II microcutting, c. microcutting transfer to Stage III rooting medium, d. rooting after 4 weeks, e. rooted acclimatized plant with new runners in greenhouse.
Michael Kane, Professor Emeritus
Environmental Horticulture Department, University of Florida
Srivas, A. and O. Sumant, 2021. Global Plant Tissue Culture Market. Allied Market Research Report