TETRAPLOID CREATION AND CONVERSIONWhat is the difference between a creation and a conversion? A "creation" generally occurs in seeds, while a "conversion" is specific to a given cultivar. Let's begin with the creation of tetraploid plants from diploid seeds.
A tetraploid plant will be created if a cell in its apical meristem (the tiny cluster of cells where all leafy growth originates) fails to segregate its chromosomes when dividing, that is, one daughter cell will receive no chromosomes (this is lethal to the daughter cell) and one receives two sets (this luckier daughter cell will survive and all other cells arising from it will be polyploid). Because the meristem is made up of more than one cell, such a plant will be a chimera, with some cells remaining diploid, and others polyploid. The plant breeder will hope that the tetraploid cells will give rise to the pollen and ovaries, because these tissues will transmit the tetraploid characteristic to their progeny.
One can chemically interfere with chromosome segregation with a chemical called colchicine. To prepare a colchicine solution, you should weigh 0.5g and dissolve into 1 liter of water. Colchicine is a potent carcinogen and should only be handled by trained laboratory personel.
A cell culture is prepared from the plant's meristem, and many plantlets are allowed to develop. In order for the treatment to be effective, plantlets must be in active vegetative growth, that is, the green apical leaves should just begin to emerge from the corms. Individual seedlings can be placed in 15 mL Sarstedt centrifuge tube, sealed and gently rocked on a mechanical rocking platform for 12-48 hours. After treatment, the solution is poured out and safely discarded, and the seedlings are rinsed in water several times, then rocked again in water for a period of time equivalent to that of the colchicine treatment. All the while, be mindful of keeping a night/day light cycle for optimal shoot health.
Seedlings will be severely weakened after treatment. Many will die from it; that is a sign that the treatment may be effective with the survivors. These weakened plantlets are best pampered in a greenhouse. Some will survive and not grow for a while. Tetraploid plants oftentimes grow more slowly than their diploid counterparts, so patience is of the essence. If you do not have access to a cytology lab or microscope to actually count the chromosomes, you can try to test your plant's pollen with diploid and tetraploid pod plants. If the pollen takes on the diploid pod plant, but not the tetraploid, you have failed. The opposite, and you have been successful.
A popular alternative is to treat the apical meristem on a growing plant in order for this plant to produce a tetraploid floral scape that will yield tetraploid pollen that can serve to fertilize other tetraploid plants.
A conversion is entirely different and more costly. It involves the physical extraction of meristematic tissue of a specific, desirable cultivar, and growing it in cell tissue culture, on shakers. The cell culture is treated with colchicine, and plantlets will be allowed to arise by stopping the shaking motion during growth. Some of these plantlets will be a tetraploid version of the diploid cultivar thus treated.
DAYLILY MUTAGENESISEthylmethane sulfonate (EMS) is a very dangerous mutagenic and carcinogenic substance that must only be used and disposed of in a laboratory familiar with the handling of powerful mutagens. Generally, seeds are allowed to soak overnight in water, and then exposed to a solution of 0.2% (w/v) EMS on a shaker for 24 hours. The seeds are then rinsed ten times, and allowed to germinate. Armed with a proven protocol, the daylily breeder should hire a professional facility to treat his seeds, and perhaps even seek their help in devising an optimal course of treatment. Successful mutagenesis is suggested by a lethality rate of 50%. Tetraploidy doubles up the chromosomes, which can be great, but EMS has the power of making point mutations, that is, it leaves the chromosomes super-structure and count largely intact, but will modify individual genes in horticulturally novel ways. It may create new shapes such as reliable polypetals, yet unseen colors and patterns, as well as totally unexpected results. You are unlikely to see anything but dominant mutations in the treated seeds. Since most mutations will be recessive, it will be necessary to self-pollinate the plants that that survived the EMS treatment. It is from their offspring that the majority of interesting new mutants will be found.
As most mutations are deleterious, a very large number of treated seeds will have to be planted in order to find the gem in the army of EMS treated runts.