A few tricks to try in wide crosses within the rose family.

In my reading about intergeneric crosses, I came across what may be tricks to try in wide crosses within the rose family.

The first is based on the following paper:

Title: Pollen tube growth and early embryogenesis in wheat x maize crosses influenced by 2,4-D

Author: Wedzony, Maria; Van Lammeren, Andre A. M.

Authors affiliation: Dep. Plant Physiology, Polish Academy Sci., Slawkowska 17, 31-016 Krakow, Poland.

Published on: Annals of Botany (London), volumn 77, pages 639-647, (1996).

Abstract: " Pollen tube growth and embryogenesis were investigated in the intergeneric Triticum aestivum times Zea mays cross. Emasculated wheat florets were pollinated with maize pollen, and at one day after pollination, the wheat plants were injected with 2,4-dichlorophenoxyacetic acid (2,4-D). The influence of 2,4-D on pollen tube behaviour was determined applying callose staining in whole mount preparations of pistils. Changes in the embryo sac and early embryogenesis were analysed on sections after various histochemical stainings. Maize pollen tubes germinated within 30 min and grew much slower through the pollen tube pathway compared with selfings of both maize and wheat. Deviations in pollen tube growth occurred such as coiling, widening and forking, irrespective of treatment with 2,4-D. Pollen tubes reached the micropyle between 5 and 24 h after pollination. 2,4-D treatment increased the number of the pollen tubes that reached the micropyle, and additionally, multiplication of sperm cells was found in the maize pollen tubes. Embryo formation was analysed at 2 d after pollination. Only when pollen tubes were in the micropyle, zygotes and embryos were observed. This points to their hybrid origin. Moreover, the embryos are likely of zygotic origin since multicellular embryos exhibited micronuclei, a sign of chromosome elimination not occurring in parthenogenic embryos and leading to haploidy of the embryos. Treatment with 2,4-D increased successful intergeneric fertilization from 18.7 to 69.3%. Immature embryos were rescued by in vitro culture from 14% of pollinated florets when excised at 14 d after pollination."

My comments: a 1 ml syringe was used to inject 1 ml of a 100 mg 2,4-D per liter of solution into the stem cavity of the highest internode. My question is: will this introducing 2,4-D result in the plant eventually dying? They removed the embryos after 14 days in order to culture in a petri dish; so, I assume the plants were still living then. If that amount of 2,4-D would eventually kill the plant, is there a chemical that could used several days after the injection of the 2,4-D in order to save the plant (such as GA3) and allow the hips to form normally?

The above (A “trick” to try with wide crosses.) should of been the title of the thread.

Second trick:

Title: Effect of high-temperature on suppression of the lethality exhibited in the intergeneric hybrid between Japanese pear (Pyrus pyrifolia Nakai) and apple (Malus

One can increase fertility by going through a callus.

Title: Tissue culture increases meiotic pairing of regenerants for barley x Canada wild rye hybrids

Author: LS Dahleen

Author affiliation: Department of Agriculture, Agricultural Research Service, Northern Crop Science Laboratory, PO Box 5677, State University Station, Fargo, ND 58105, USA.

Published in: The Journal of Heredity, volumn 90, pages 265-269, (1999

Abstract: “Canada wild rye (CWR; Elymus canadensis L.) expresses traits such as barley yellow dwarf virus resistance, winter hardiness, and drought resistance. Hybrids between CWR and barley (Hordeum vulgare L.) are sterile, precluding transfer of these traits into barley. Callus cultures were initiated from these hybrids to promote chromosome recombination and possibly restore fertility. The objectives of this study were to determine the effects of tissue culture and cytoplasms on meiotic pairing of 17 intergeneric hybrids generated by crossing Betzes barley and CWR, and 117 regenerants, and to examine meiosis in a BC1 plant from a regenerant x barley cross. Meiotic abnormalities were common, including chromosome bridges, lagging univalents, and micronuclei. Regenerants with 2n = 21, 20, or 22 chromosomes had significantly greater pairing and fewer micronuclei than the hybrids (2n = 21). Regenerants with partially or completely doubled chromosome complements (2n = 36-42) showed chromosome instability. Hybrids and regenerants with barley cytoplasm had significantly more pairing and fewer micronuclei than those with CWR cytoplasm. Application of barley pollen to florets of a stably doubled (2n = 42) regenerant resulted in a single BC1 plant that survived to develop spikes. This plant had the expected 28 mitotic chromosomes and showed meiotic chromosome number instability. Backcrosses to this plant produced small embryos for rescue, indicating that the BC1 plant is partially female fertile. Attempts to recover BC2 plants are in progress. Results indicate that tissue culture significantly increased chromosome pairing, providing additional opportunity for genetic recombination between the two species.”

My comment: The body of the article suggests doing the cross both ways as one way may be more admendable to the procedure.

My second comment: a callus can also be used to slightly modify a plant. For example: if you create a callus of William Baffin and then regenerate plants from the callus; some of the plants will be slightly different than William Baffin - one could be shorter growing, one could have a different color or size flower, etc. I had posted on this aspect in the past. If someone is interested in rose literature concerning this aspect of calus regeneration, let me know and I will try to find my previous information.

Link: jhered.oupjournals.org/cgi/content/abstract/90/2/265?maxtoshow=&HITS=10&hits=10&RESULTFORMAT=&author1=dahleen%2C+ls&andorexacttitle=and&andorexacttitleabs=and&andorexactfulltext=and&searchid=1073529781343_382&stored_search=&FIRSTINDEX=0&sortspec=re

Well, I did get e-mail requests for further information about using calli.

I interpret this paper as suggesting that if you want to develop a new quality rose quickly, you should utilize the best rose that you can obtain and use tissue culture to develop calli from it. When you regenerate plants from the calli, you should have many quality mutants - more or less petals, different colors, dwarf growth habit, etc (this is not immediately obvious from the abstract, one has to read the full paper). For example, lets say you are impressed with the disease resistance and winter hardiness of William Baffin but are “turned off” by its ten foot height. By using this procedure you could select for dwarf growth habit. I assume that you could also introduce any color sports, petal sports, etc.

Title: A comparison of the somaclonal variation level of Rosa hybrida L. cv Meirutral plants regenerated from callus or direct induction from different vegetative and embryonic tissues

Authors: Laurence Arene, Carole Pellegrino & Serge Gudin

Authors affiliation: Meilland, Domaine de Saint Andre, 83340 Le Cannet des Moures, France

Published in: Euphytica volumn 71, pages 83-90, (1993).

Abstract: “Flowering plants of Rosa hybrida L. cv Meirutral have been obtained either from direct regeneration of adventitious shoots on leaf and root fragments, or through organogenesis and somatic embryogenesis on calli derived from anther, ovule, petal, sepal, receptacle, leaf. stem internode. root and zygotic embryo tissues. The calli derived from floral parts exhibited rhizogenesis. In this case direct induction of adventitious shoots from selected roots had to be performed in order to generate plants. A histological study of the morphogenetic calli was carried out. The plants regenerated directly and those regenerated from calli of leaf, stem internode, root and zygotic embryo tissues, together with reference plants propagated by cuttings, were compared on a phenotypic basis by taking into account petal number, form and color, and plant growth habit. From these observations, it can be concluded that directly regenerated plants are as stable as reference plants while plants regenerated from callus are unstable, especially those derived from zygotic embryo tissues.”

The following is from the body of the paper:

" Upon first blooming in the greenhouse, no phenotypic variants were observed with plants produced from either cutting propagation (reference plants) or direct regeneration from leaves and roots. On the contrary 21.7 and 70% of variation could be observed with plants regenerated from calli of vegetative parts and zygotic embryos, respectively. The recorded variations remained stable and were still observed on 2 successive flushes occurring on plants propagated by cuttings taken on the variants detected during first flush of regenerated plants (5 cuttings per variant). Among the 152 recorded variant plants, 83 significantly differed from the original cultivar by their number of petals (Fig. 5), 16 by their dwarf growth habit (Fig. 6), 4 by their round shaped petals (Fig. 7),12 by their color, 33 by both their growth habit and number of petals, 4 by both their petal number and color. The variant lot differing by the number of petals was represented by 3 plants with 71 + 5 petals, 8 plants with 25 + 3 petals. 36 plants with 20 + 2 petals, 30 plants with 10 + 2 petals, and 6 plants with 5 petals. The lot differing by the petal color was represented by 6 plants with orange petals. 3 plants with indian pink petals and 3 plants with dark red petals. The 33 plants that differed by both their dwarfness and number of petals all had 10 petals. The 4 plants that differed by both their petal number and color all had 5 orange petals."

The final statement in the DISCUSSION SECTION is: “Regenerating plants from calli might be attractive to the breeders as the somaclonal variation generated by this system is known to be higher than that generated by other systems such as chemically induced mutagenesis (Gavazzi et al., 1987).”

Euphytica 71: 83-90,1993 .
A comparison of the somaclonal variation level of Rosa hybrida L. cv Meirutral plants regenerated from callus or direct induction from different vegetative and embryonic tissues
Laurence Arene, Carole Pellegrino & Serge Gudin

I came across this paper again yesterday, but read it with different understanding.

For example, the authors include a very interesting picture on page 6. The original type is shown in the middle. To the left is a semi-double, and to the right is one that is more double than the original. Of course, such results could not be expected for every cultivar, but maybe that almost-perfect seedling could be made just a bit more double.

Or, perhaps some of those too-too-double old roses that refuse to produce even one style or stamen might be nudged towards just a little fertility. I’m thinking of ‘Rosette Delizy’ and ‘Mme Hardy’, to name just two.

And on the related subject of sterile hybrids, Larkin (1998) gave some suggestions towards the use of tissue cultures for getting those foreign genes introgressed. He cites the example of Thinopyrum intermedium, a grass that will cross with wheat, but whose chromosomes do not pair. A hybrid was maintained as a tissue culture, and eventually the researchers had wheat with enough Thinopyrum DNA mingled in to give immunity to Barley Yellow Dwarf Virus.

Has anyone seen an apple tree with blackspot or a hawthorn with RRD? They might be suitable donors.