Plant Tissue Culture Micropropagation Is Growing

Plant Tissue Culture which has been around for decades, is a way to reproduce new plants from the mother tissue and is used as an alternative to cloning.  It originated as a solution for hard to germinate orchids, but has been “sprouting” this new standard throughout the cannabis community and shows great promise for high production farming.

More specifically, plant tissue culture is a collection of techniques used to maintain or grow plant cells or tissues under sterile and virus free conditions and growing them with a nutrient aid such as ORIGINAL CHO ORANGE FORMULA — A bio-based clone and seed treatment to buffer pH.

Benefits Of Micropropagation
Plant tissue culture is widely used to produce clones of a plant and this technique is known as micropropagation. Different techniques in plant tissue culture may offer certain advantages over traditional methods of propagation, including:
The production of exact copies of plants that produce particularly good flowers, fruits, or have other desirable traits.
To quickly produce mature plants.
The production of multiples of plants in the absence of seeds or necessary pollinators to produce seeds.
The regeneration of whole plants from plant cells that have been genetically modified.
The production of plants in sterile containers that allows them to be moved with greatly reduced chances of transmitting diseases, pests, and pathogens.
The production of plants from seeds that otherwise have very low chances of germinating and growing.
To clear particular plants of viral and other infections and to quickly multiply these plants as ‘cleaned stock’ for horticulture and agriculture.
Plant tissue culture relies on the fact that many plant cells have the ability to regenerate a whole plant (totipotency). Single cells, plant cells without cell walls (protoplasts), pieces of leaves, stems or roots can often be used to generate a new plant on culture media given the required nutrients and plant hormones.

Overview Steps In The Process
Preparation of plant tissue for tissue culture is performed under aseptic conditions under HEPA filteredair.  The tissue is then grown in sterile containers, such as petri dishes or flasks in a growth room with controlled temperature and light intensity. This is because living plant materials from the environment are naturally contaminated on their surfaces (and sometimes interiors) with microorganisms, so their surfaces are sterilized in chemical solutions (usually alcohol and sodium or calcium hypochlorite) before suitable explants are taken. The sterile explants are then usually placed on the surface of a sterile solid culture medium, but are sometimes placed directly into a sterile liquid medi um, particularly when cell suspension cultures are desired. Solid and liquid media are generally composed of inorganic salts plus a few organic nutrients, vitamins and plant hormones. Solid media are prepared from liquid media with the addition of a gelling agent, usually purified agar.
The composition of the medium, particularly the plant hormones and the nitrogen source (nitrate versus ammonium salts or amino acids) have profound effects on the morphology of the tissues that grow from the initial explant. For example, an excess of auxin will often result in a proliferation of roots, while an excess of cytokinin may yield shoots. As cultures grow, pieces are typically sliced off and subcultured onto new media to allow for growth or to alter the morphology of the culture.
As shoots emerge from a culture, they may be sliced off and rooted with auxin to produce plantlets which, when mature, can be transferred to potting soil for further growth in the greenhouse as normal plants.

Micropropagation VS Traditional Cloning
The cost benefits become evident when comparing micropropagation to traditional cloning or seed propagation. A traditional indoor farming operation requires care that is costly:  daily watering and fertilizer, electricity for LED lights and climate control, and space required to house the plants. When you’re talking about a large scale commercial operation, the prices climb steadily. Micropropagation cultures, however, are surprisingly low maintenance.  They need to be divided and transferred to a new grow medium every 4 to 6 weeks.  The only maintenance is periodic observation to be sure all is going as planned.
Micropropagation also allows for the generation of thousands of plants in a very short time period with very small square footage, something that would be impractical with traditional cloning techniques.  To help put this in perspective, think about this:  If properly cared for, one explant will multiply indefinitely.
Pros and Cons
Space saver, much less storage is required to preserve genetics
Provides exact replicas of the mother plant, creates uniform offspring
Sterile Environment reduces risk of pests and disease
Speeds up the breeding / pheno-hunting process
Able to produce an endless amount of plants from one “cutting” (explant).
Minimal daily care
Allows for year-round propagation
Steep build-out investment for a large-scale operation
Requires patience, takes more time than traditional cloning if conducted on a small scale
Requires extreme attention to detail
Sterile/controlled environment is necessary

Micropropagation could be an alternative to cloning in the future, but implementing tissue culture labs at every farm is somewhat impractical. If you look at the fruit tree industry, that is not what is happening. Large-scale fruit tree cultivators often source their micropropagated clones from an experienced nursery. As the cannabis industry expands and becomes integrated throughout the states, farms are likely to follow a similar model.
Details In Getting Started
However in the meantime, cannabis farms are establishing personal tissue culture labs left and right. So what about the logistics of creating your very own lab-grade micropropagation lab.
Required Equipment:
– Laminar Flow Hood
– HEPA Filter
– Magnetic stirrer
– pH measure
– Glass measuring gradual
– Culture bottle with lid
– Dissection tools (Forceps, spatula, scalpel, tweezers, scissors)
– Conical flasks
– Refrigerator
– Microwave
– Pipette measuring
– Shaker
– Aluminum foil
– Disinfectants (Ethanol, Clorox, Tween 20)
– Bunsen burner (to sterilize equipment)
The first step of micropropagation process is prepping the media. The media used during the first stage of micropropagation is a nutrient-rich substrate filled with chemical compounds designed for growing cultures, essentially food for the plant tissue. Recipes and techniques are constantly changing as scientists experiment and adapt to what suits plant cultures best.
Pre-mixed medias are available to purchase online, which I recommend. You can always make your own but the recipe calls for a variety of uncommon nutrients. For those who are curious, here’s what goes into the average micropropagation media:
– Macronutrients: NH4NO3, KNO3, CaCl2.2H2O, MgSO4 7H2O, KH2PO4
– Gamborg’s B5 Vitamins: Myo-inositol, Nicotinic Acid, Pyridoxine, Thiamine HCI
– Micronutrients: H3BO3, MnSO4.H2O, ZnSO4.7H2O, KI, Na2MoO4.2H2O, CuSO4.5H2O, CoCl2.6H2O
– Sucrose
– Agar
– Coconut water (optional)
– Distilled Water
– The pH is then adjusted to 5.8 using hydrochloric acid or sodium hydroxide.
The next step is prepping the explant (plant material). Freshly sprouted nodes, the newest growth on the mother plant, is the best source material because it hasn’t had a chance to be exposed to diseases or pathogens. Once the cutting has been taken, the plant material needs to be heavily disinfected or else we risk the growth of unwanted cultures. To clean, simply place your explant in a test tube with ethanol, swish around for a couple minutes, drain, and repeat the process one more time. Discard the ethanol and rinse your plant material (still in the test tube) with distilled water. Discard the water and leave the plant material in a laminar flow hood to dry.
Once the explant is prepped, it’s time to sterilize the work station and all material with ethanol. This step requires a conical flask and forceps which both need to be sterilized. Once the conical flask is filled with the media, the explant is transferred to the flask and sealed with a cap or covered with aluminum foil.
Flasks need to be kept in a mild-temperature environment with plenty of light. And before you know it, infant seedlings will begin to form in a couple weeks!
After a month or two, depending on the plant, the seedling will be noticeably developed and ready for transplant.Since the seedlings were created in such a sterile, controlled environment, it’s important not to shock them during transplant. A small terrarium or greenhouse environment is recommended as humidity and temperature should not fluctuate for the first week. You can make your own humidity dome with a plastic bag placed over the individual pot — poke a few holes in the bag and spray with water and voila, you have a humidity chamber.
Over the next couple of weeks, slowly harden off the plantlet by gradually exposing it to a more dry and bright environment.
As long as the media is kept fresh and changed every 4-6 weeks, plant genetics can be preserved via plant tissue culture for many years.
This technology has the potential to produce plants unlike anything produced through conventional cloning. The process of culturing cannabis rejuvenates old, mature plant tissue into a healthy, vigorously growing juvenile state, in some ways like newborn seedlings. This plant fountain of youth gives cultured plants many new advantages for growers: higher yields, stronger growth, and better resilience to environmental stress. Plus, mass production of high quality disease free clones mean greater availability and variety for growers.  I hope this information shows you that getting started with plant tissue culture isn’t as intimidating as it’s made out to be.

Source: (12/12/17)