Plant tissue culture processes were initially developed and practiced in university and government-based laboratories. In recent years, however, the process has moved beyond these research facilities to widespread use among commercial enterprises as a cost-effective tool for plant propagation, new variety introductions and research.
Plant tissue culture has revolutionized the flower and nursery markets by making valuable new hybrid clones available in commercial quantities comparatively soon after their first discovery. It is possible to start with a single plantlet and, in the space of 10-12 months, create in excess of 250,000 identical copies or clones.
Plant Propagation Methods
Plants produced by vegetative propagation – including top cutting, root divisions, pseudobulbs, offshoots and plantlets – are genetically identical to the mother plant and thus members of a single clone. Plant tissue culture is a laboratory-based extension of these plant propagation techniques.
Through tissue culture, very large numbers of identical plantlets can be derived from one mother plantlet. This technology and the resulting plantlets now form the basis of many plant nursery and flower trade industries.
The Tissue Culture Process
The mother plant selected should be healthy and free from all microorganisms. A piece of live tissue is removed from a part of the plant and placed into culturing flasks containing appropriate nutrient media under aseptic conditions.
In successful culture, these cells will divide, multiply and differentiate into thousands of plantlets having the same characteristics as the parent plants. In tissue culture, the presence of bacteria and fungi will prevent the growth of the plant tissue. It is important that the plant material used is thoroughly sterilized and the procedure is carried out in an aseptic environment.
Laboratory techniques and specialized equipment such as a laminar flow cabinet combine to present an area for the manipulation of sterile plant tissues. A working environment that has virtually all of the bacteria and fungal spores removed is a requirement for successful plant tissue culture. A sterile environment is prepared, now we prepare sterile instruments to remove an apical shoot from a plant.
Tissue Culture Media
The excised bud is transferred into a tube containing a sterile nutrient medium. The success of tissue culture depends very much on the stage of explant selected, the sterilization period and the type of culture media used; different types of plants require different sets of culture media. The rich tissue culture media provides a good food source for bacteria and fungi; therefore, precautions against microbial contamination must be taken in all in vitro procedures.
Once a plant has been successfully entered to in vitro culture and is growing, a multiplication program may be carried out.
Careful manipulation of the culture media and the plant tissues will yield a 4-10 fold increase in plant numbers every 18-40 days.
When sufficient plant numbers have been developed, a process to move the plantlets from the confines of the culture vessel to a greenhouse is undertaken. The following table compares a conventional propagation scheme on the left with a plant tissue culture production system on the right. Note the number of days and the number of resulting plants:
|Tissue Culture Propagation|
|Day 1||One plant in tissue culture media|
|Day 40||5 plantlets cut & transplanted in media|
|Day 80||25 plantlets|
|Day 120||125 plantlets|
|Day 160||625 plantlets|
|Day 200||3,125 plantlets|
|Day 240||15,625 plantlets|
|Day 280||Transplant 15,625 plants to Greenhouse|
|Day 320||Transplant 15,625 plants to Field|