QBARS - v31n4 Using Colchicine for Plant Improvement

Using Colchicine For Plant Improvement
Ellis A. Jones, Eugene, Oregon
From a paper given at the 1977 Breeders Roundtable in Eugene, Oregon

In 1966 Dr. August Kehr, gave a detailed report in the ARS Bulletin on the possible procedure for obtaining tetraploidy in rhododendrons. In 1968 he kindly furnished me with an Agricultural research paper by Prior & Frazier on colchicine-induced, tetraploid, evergreen florists azaleas. This paper detailed the procedure which they used in obtaining tetraploid evergreen azaleas in 2 treatments. In one treatment 12% of the plants were effected and, in the other, 22% of the plants showed the effects of treatment.
The measure they used to indicate the increases in the size of the plant parts was to measure the pollen grain size between untreated plants and those which were effected by colchicine. The average increase in pollen grain size was 35.4% with three clones increasing size by approximately 50%. It is my assumption that the increase in size of the pollen grains would indicate the relative increases in thickness and size of all of the other plant parts.
In their treatments, Prior and Frazier removed the terminal tip and treated the next four lower auxiliary buds for three days thereafter with treatments on alternate succeeding days for six days. They used a 0.5% solution of colchicine in a 10% glycerin solution plus a spreader sticker. I followed the guidelines of Dr. Kehr and the Agricultural Research paper with some deviation.
Firstly, I potted up single-stem, rooted cuttings in four inch pots. These were forced into fast growth under greenhouse conditions with 24 hour fluorescent light, using deluxe, warm-white bulbs. I then amputated the stem above the highest auxiliary bud and started treating the top four buds three or four times per day. In actuality, they were practically under continuous contact with the liquid colchicine treatment and the greenhouse was very humid and most mornings the buds would still be wet from the last treatment of the previous day. I continued treatments daily until I saw distorted leaves from the new growth. I forced growth from only the treated buds by rubbing lower emerging buds as they appeared. I stopped treatment only when I could see some visual effect upon the new growth. Treatment on some clones continued for up to 28 days.
I treated small leaved scaly rhodies such as R. impeditum , 'Ramapo', 'Dora Amateis', 'Windbeam' and the white form of R. carolinianum , R. vaseyi 'White Find', 'Pioneer', and 'P. J. M.', about 60 plants, as I recall. I have flowered three plants in which the chromosomes were doubled, for three or four years to date. This is a success ratio of about 5%.
I want to emphasize, that I had no special training and no special facilities. I used a very small greenhouse with the plants on one little bench under an eight foot fluorescent fixture. Any one of you here in this room can easily provide the meager facilities and conditions necessary to induce chromosome doubling. I would hope that some of you would make the attempt (this is my primary reason for being here today) to encourage you to double the number of chromosomes of some of our present clones.
One of the plants which I doubled was the white form of R. carolinianum . As you are undoubtedly aware, Dr. Kehr also produced a doubled white carolinianum , I believe being the first to accomplish doubling in rhododendrons, but the method of treatment was with treatment of colchicine upon germinating seeds. I prefer the terminal tip method simply because you don't have to live so long to see them flower and you have a before and after affect - you make your evaluation as to benefits derived. And I want to say that each spring, as I look at the plant of white carolinianum along side of the original form, it is to me a miracle. The flowers, and foliage are completely superior and the original form has become last year's model, it is obsolete. But of more significance is the fact that there is an entirely new and hardy gene pool available for crossing into divergent, scaly-leaved rhododendrons having 52 chromosomes which may be less hardy.
Now I want to mention an apparent failure. A year or so later I did approximately the same things, I took ten larger plants of some 30 large-leafed clones which were in two gallon containers. I placed these outside in a lath house under incandescent light. This was in July with the weather being warm. I could see the effects of colchicine on these plants but probably not to the same extent which I saw it in the greenhouse. I have probably flowered 20% of these without any results and am not at all optimistic.
I believe I made a fundamental mistake when I treated the plants outside. While the days were hot, the nighttime temperature may have dropped to 60°F. It is my understanding that chemical reactions practically double with each ten degrees of temperature increase so that I probably was not getting fast cell division and proper chemical reactions during the cooler night hours. I tell you of this experience so one of you won't repeat the error. Now I also was successful in doubling the number of chromosomes of two plants of rhododendron 'Windbeam'. The flower size increase is proportionate to the slide which you saw of the tetraploid carolinianum album and the doubled plant is certainly superior to 'Windbeam' itself. But here, a fortunate circumstance occurred which makes the tetraploid form of 'Windbeam' genetically valuable.
'Windbeam' was an open-pollinated seedling from a plant of R. racemosum times carolinianum . It also is a mule (it is sterile) and does not produce seed. I can now tell you that the original sterile 'Windbeam' has 39 chromosomes instead of the usual diploid 26 and is a triploid. When the number of chromosomes was increased, the increase was from 39 chromosomes to 78 chromosomes. I haven't proved this with an electron microscope but I base the theory on these facts. If it were a diploid the doubled form would cross with other scaly 52 chromosome forms as does the doubled form of white carolinianum . It does not produce seed from 52 chromosomes crosses but does produce seed when crossed with 78 chromosome crosses such as R. davidsonianum , rubiginosum , oreotrephes , yunnanense , and some others, and should be a real candidate for crosses with flavidum , and the Cinnabarinum, and Maddenii Series.
O. K. you say. Why double chromosomes and produce a new race of larger flowered monsters?
Well, I can tell you that if I offer one of my retail customers a plant of 'Blue Peter' with normal size trusses and one with larger trusses, I know which one he's going out the door with.
Also, I can remember, from someplace in the literature, when some plant society, day lilies, iris, or whatever, voted on their favorite flower varieties in order, and they ran chromosome counts of the favorites about 90% turned out to be naturally produced tetraploids.
One of the advantages of tetraploidy would be that, because of thicker flower petals you would get increased frost resistance. Those of you who have flowered seedlings and gone out in the field to look at them, know that flowers without good substance turn to mush from the frost, while adjacent plants are not damaged. Colchicine will give more flower substance.
Let's consider 'Crest'. If you doubled 'Crest' the thickness of the flower petals would increase. Visually, this additional thickness might very well give you the illusion of deeper flower intensity.
Would you like a plant of 'Crest' with much larger flowers, greater substance and an apparently deeper color? It will only take a few drops of colchicine and I urge each of you to have a go at it.