An amphidiploid of Rosa banksiae and Rosa laevigata induced by colchicine
Robert E. Basye
P.O. Box 494 Caldwell, Texas 77836

According to Rehder, most wild roses belong to the subgenus Eurosa of the genus Rosa. This subgenus is divided into ten sections, two of which are Banksianae and Laevigatae. The first of these contains only R. banksiae and the related R. cymosa. The second contains only R. laevigata. It might be suspected, therefore, that R. banksiae and R. laevigata do not hybridize easily with most other roses, and this is indeed the case. Being native to southern China, they are also not very hardy. With these shortcomings, it might be thought useless to try and bring them into the society of modern roses, even though they have some very sterling virtues such as evergreen foliage, longevity, and disease resistance. But let us remember that the tea and china roses, though tender, have contributed the ever-blooming trait to our modern roses. Who among us would now sacrifice this treasured feature just because modern roses are not as hardy as we would like?

There is yet another difficulty. Both R. banksiae and R. laevigata are diploids with 14 chromosomes, while most garden roses are tetraploids with 28 chromosomes. The cross, R. banksiae x R. laevigata, makes a fine and healthy plant, but it is almost completely sterile, and of course, is still a diploid. By applying colchicine to this cross, however, we can make it both highly self-fertile and tetraploid! The magic behind this use of colchicine is perhaps the greatest contribution to genetics and plant breeding in the present century.

Colchicine is an alkaloid found in the corms and seeds of the autumn-blooming crocus, Colchicum autumnale, native to regions near the Mediterranean Sea and the Black Sea. It flowers in the autumn, but waits until spring to produce leaves and fruit. Some effects of the drug were known in ancient times. For example, it was used as early as the sixth century A.D. to relieve the acute pain of gout. Physicians still use it today for this purpose.

In the 1930s, botanists began to notice the effects of colchicine on cell division. Then, in 1937, the first doubling of chromosomes in plant cells was reported. (A. Blakeslee and A. Avery, Methods of Inducing Doubling of Chromosomes in Plants, Journal of Heredity, Vol. 28,1937: 393-411. This discovery led to a veritable torrent of interest and research in genetics and plant breeding which persists to this day.

Most of this research, however, has been devoted to the food and fibre crops, while ornamental plants (yes, even the rose) have been sadly neglected. To the best of my knowledge, the rose amphidiploid described here is the first to be obtained with colchicine. Nature has her own way of producing them on rare occasions by uniting two unreduced gametes, as we think happened centuries ago with cotton, coffee, and tobacco. But to verify such a production and rule out the involvement of a third species is not easy. That is why I had to speak of a "probable amphidiploid" in the 1987 American Rose Annual.

Many techniques for applying colchicine to double the chromosomes of plants have been tried. I present here, in careful detail, essentially the method which was suggested to me 28 years ago by the British rosarian and taxonomist, Gordon D. Rowley, and which I have found quite effective through the years in producing both autoploids and alloploids. The method is effective mainly with dicotyledons, which include roses. We describe the production of an alloploid, that is, a plant which results from crossing two species and then doubling the chromosomes of the cross. In our case, the two species are both diploids, so the alloploid can also be called, more specifically, an amphidiploid.
We begin in the greenhouse with a flat of germinating seeds of the cross, R. banksiae x R. laevigata. I must not digress here to describe the many details leading up to this point; although, the prime condition of the seedlings is of prime importance to success. I will just mention one thing. The seeds of this cross rarely germinate the first spring after harvest and must be held over till the following year. This might be expected, since seeds of both parents behave in the same way.

I should also mention that the seed parent, R. banksiae, is a completely thornless plant which I raised from seed sent to me many years ago by the Italian rose breeder, Domenico Aicardi. A mislabeled picture of it is shown on page 125 of the 1988 American Rose Annual.

The medium in which the seeds are sowed is one part peat moss, one part milled sphagnum moss, and two or three parts thoroughly decomposed leafmold, all finely screened. The young seedlings, when they appear, are easily plucked out of this soft medium with a pair of tweezers and placed in water before being potted. The ideal time is when the cotyledons have emerged, but the neck has not yet straightened out. Plucking the seedlings a day on either side of this time should also be satisfactory.

Without being too specific, my potting soil contains 1) rich soil from the floodplains of a nearby river, 2) some peat moss, 3) a little sand, 4) a generous amount of leafmold, 5) a little gypsum, 6) a little 20-percent superphosphate, and 7) a little slow-acting fertilizer such as Magamp or Osmocote. To pot the seedlings, fill 2-inch clay pots with moderately moistened soil and firm level with the rim. Water the soil until saturated. With a paring knife, open a slit in the center of each pot and insert the seedlings. After another watering, they may be moved into the greenhouse near sundown.

Within a day or two, the little seedlings will be erect and reaching for he sun, and the two cotyledons will become greener, larger, and plumper. Then, in the crotch between the cotyledons, the second leaves will become just barely visible. With cells dividing rapidly, this is the time to apply the colchicine to the tiny meristem. I use 0.5-percent aqueous colchicine, prepared by adding one-half gram of colchicine to 100 cubic centimeters of water. It is best applied with a medicine dropper and refrigerated in a small bottle when not in use.

With the medicine dropper apply, in late afternoon, a drop or less to cover the tiny second leaves in the crotch. How do you apply less than a drop? This is done by touching the end of the dropper to the crotch, pressing out the desired fraction of a drop, then pulling away the dropper before releasing pressure. Just enough colchicine to cover the tiny meristem region through the night is required. Using half a drop doubles the number of seedlings that can be treated with the precious fluid. In early morning, the seedlings are sprayed with water to wash away the colchicine. This treatment is carried out on four successive nights, after which the treated seedlings receive only normal greenhouse treatment.

To prevent the colchicine from evaporating overnight, polyethylene or plastic covers can be designed to span the greenhouse bench. Or the seedlings may be placed in a tight cold frame or in plastic boxes with covers.
In early spring, the treated seedlings are moved outside to the nursery where they are mulched and watered through the summer. In early autumn they are examined for signs of chromosome doubling. Even before the plants have bloomed, evidence is found in the gigas characteristics of mature foliage. The leaflets are usually a little larger and thicker than comparable diploid leaflets not affected by the colchicine. This comparison of thickness is made by simply feeling the leaflets between the thumb and forefinger. Leaflets with doubled chromosomes may also be a little darker green and have a crinkled appearance. The midrib of the leaf is usually a little broader and sometimes shows a little tortuosity.

In early autumn of 1985, I tagged three of the crosses as being likely candidates for having a tetraploid branch. One of these plants died that winter, but the other two are thriving today-three plants of each having been budded off onto the understock ‘Fortuniana’. I focus on one of these, bearing #86-3, as being distinctly superior to the other.
A microscopic search showed only tetraploid cells in the meristem tissue of the selected branch of #86-3, suggesting that this entire branch might have grown from a single, colchicine-doubled cell in the tiny original seedling. If this is true, it suggests that the above technique for using colchicine may be valuable for avoiding chimeras.
Number 86-3 and its parents are the first roses of spring to bloom. They are, therefore, subject to damage by late freezes. Such was the case in 1988 when #86-3 produced only a dozen hips. But in the autumn of 1989, as I write this, the three big bushes are loaded with fruit, supporting the rule that the more sterile the F1 cross, the more self-fertile will be the associated amphidiploid.

Many of the hips now ripening are selfs which I made in the spring. It is my first aim to acquire, and make homozygous, the thornless feature of the R. banksiae parent. Meanwhile, though, I could not resist making cross, #86-3 x Cyt.67, where the pollen parent is a tetraploid form of laevigata which I obtained with colchicine in 1965. Many hips of this cross are now ripening. R. laevigata ('Cherokee Rose') has the distinction of bearing a large and beautiful, pure white flower, always one to a stem. In distinct contrast, R. banksiae blooms in many-flowered umbels of small roses.

I must mention one other cross that I made, R. carolina alba x #86-3, whose hips are now ripening. I described in the 1986 American Rose Annual how R. carolina alba was effective in breaking down the barrier between R. laevigata and R. bracteata. I am hoping that R. carolina alba will eventually aid in breaking down many barriers between #86-3 and other roses. It should also improve hardiness since it was native to Maine. I will be glad to send budwood of #86-3 to rose breeders who live in the South and have understocks. You need only let me know when your understocks are receptive.

Transcribed from the 1990 American Rose Society Annual.