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June Liu, Zhimin Yang, Weiling Li, Jingjin Yu, and Bingru Huang

cold tolerance in seashore paspalum through traditional breeding efforts as a result of insufficient genetic variability in cold tolerance or the lack of cold-tolerant cultivated germplasm ( Duncan and Carrow, 1999 ). Somaclonal selection is a powerful

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Chang-Yeon Yu and John Masiunas

The objective of this study was to investigate the chromosomal and genotypic variation in regenerated plants of Solarium and Lycopersicon. Calli of Lycopersicon peruvianum genotypes PI199380, PI126345, PI251301, and LA1373, along with Solanum ptycanthum were transferred onto media consisting of MS salts with Gamborg vitamins. The shoots formed were rooted in vitro and transferred to greenhouse soil. Actively growing root tips were harvested and pretreated, fixed, hydrolyses and stained. Pollen mother cells were fixed in propionic alcohol solution and stained with aceto-carmine. The number of chromosomes were counted. The greatest variation was in Solanum ptycanthum with chromosome numbers ranging from 18 to 60 (2n=24). Progeny analysis for 12 somaclones of Solarium ptycanthum was done by selfing for two generations. Morphology, shoot height, and weight were determined in each generation. The amount of variation differed among the somaclonal lines.

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Brian W. Trader*, Hope A. Gruszewski, Norman I. Barclift, Richard E. Veilleux, and Holly L. Scoggins

Coreopsis grandiflora (tickseed) regenerates readily from leaf segments allowing the possibility to exploit somaclonal variation as a means to develop novel phenotypes. We used true leaves from in vitro seedlings of Coreopsis grandiflora `Domino' grown on MS basal medium as explants in a series of experiments to evaluate the effect of media, leaf explant orientation, and genotype on shoot regeneration. Genotype accounted for most of the variation with two particular seedlings regenerating freely and eight others generally recalcitrant. From these two seedlings, designated E2 and H2, shoots were regenerated and acclimated to the greenhouse over a period of weeks. Once the plants had established (≈6 weeks after acclimatization) they were vernalized by moving them to a lighted bench (12-h photoperiod) in a walk-in cooler at 4 °C .On transfer back into the greenhouse, the plants flowered within a few weeks; 15 of 175 somaclones were selected based on distinct differences in flower orientation and appearance. The selected somaclones were propagated by division and transplanted to the field in May 2002 in a randomized complete-block design with three-plant plots and three replications, to determine if the novel characteristics persisted through an additional propagation cycle. In the field, plant height, leaf dimension, flowering, and flower dimensions were scored in June-July 2003. Significant differences were found between somaclones and the original E2 and H2 similarly propagated seedlings for desirable (more petals per flower, greater flowering, shorter plants), undesirable (less flowering, smaller flowers), and neutral (narrower leaves, taller plants) traits.

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Jinggui Fang, Chih Cheng Chao, Richard J. Henny, and Jianjun Chen

Plant tissue culture can induce a variety of genetic and epigenetic changes in regenerated plantlets, a phenomenon known as somaclonal variation. Such variation has been widely used in the ornamental foliage plant industry as a source for selection of new cultivars. In ornamental aroids alone, at least 63 somaclonal-derived cultivars have been released. In addition to morphological differences, many somaclonal aroid cultivars can be distinguished by amplified fragment length polymorphism (AFLP) analysis. However, a few cultivars have no detectable polymorphisms with their parents or close relatives by AFLP fingerprints. It is postulated that DNA methylation may be involved in the morphological changes of these cultivars. In this study, methylation-sensitive amplification polymorphism (MSAP) technique was used to study DNA methylation in selected somaclonal cultivars of Alocasia, Aglaonema, Anthurium, Dieffenbachia, Philodendron, and Syngonium. Results showed that polymorphisms were detected in the somaclonal cultivars, suggesting that DNA methylation polymorphisms may associate with tissue culture-induced mutation in ornamental aroids. This is the first study of methylation variation in somaclonal variants of ornamental foliage plants. The results clearly demonstrate that the MSAP technique is highly efficient in detecting DNA methylation events in somaclonal-derived cultivars.

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Mohammad Sadat-Hosseini, Kourosh Vahdati, and Charles A. Leslie

regenerated from somatic embryos could exhibit somaclonal variation ( Bradaï et al., 2016 ). Heritable adverse changes due to somaclonal variation can be a problematic in tree-breeding programs, which typically encounter long juvenile phases before phenotypic

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Mark Bridgen

The potential value of somaclonal variation for economically important plants is well-documented. The process of somaclonal variation can arise from a controlled or a random source of variation. Variability can be obtained by applying cellular pressures and selection. Valuable resistance to diseases and nematodes has already been accomplished with somaclonal variation; now, plant tolerance to pests has been realized. Tetranychus urticae, the two-spotted spidermite, and Trialeurodes vaporariorum, the greenhouse whitefly, were disinfected and introduced to aseptic shoot cultures of Torenia fournieri. These pests were allowed to feed until such time that their populations decreased due to the absence of food. The plant cells that remained after feeding were induced to form adventitious shoots and plantlets. These regenerated plantlets were acclimated to greenhouse conditions and evaluated for tolerance to the pest to which they were subjected in vitro. Highly significant differences were found in somaclones for both the two-spotted spidermite and greenhouse whitefly when compared to control plants. A wide range of variability was observed among the somaclonal population. There were significantly fewer mite eggs laid on plants regenerated from in vitro cultures screened with two-spotted spidermites than on seed-sown controls. Regenerants from cultures screened with whiteflies in vitro had fewer eggs, immatures and live adults than controls. The potential for somaclonal variation to be used as a method to develop insect resistant plants will be discussed.

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Brian W. Trader, Hope A. Gruszewski, Holly L. Scoggins, and Richard E. Veilleux

Coreopsis species (tickseed) can be regenerated from leaf segments allowing the possibility to exploit somaclonal variation as a means to develop novel phenotypes. We used true leaf explants from in vitro seedlings of perennial C. grandiflora (A. Gray) Sherff `Domino' and `Sunray' grown on Murashige and Skoog (MS) basal medium. Two of ten seedlings of `Domino' regenerated freely and others were generally recalcitrant. From these two seedlings, designated E2 and H2, shoots were regenerated and acclimatized to the greenhouse. About 175 plants were established and vernalized from which somaclones were selected based on distinct differences in flower orientation and appearance. The selected somaclones were propagated by division and transplanted to the field in August 2001 in a randomized complete block design with three-plant plots and three replications to determine whether novel characteristics persisted through an additional propagation cycle. In the field, plant height, leaf dimension, flowering, and flower dimensions were scored in June and July 2003. Differences were found between somaclones and similarly propagated E2 and H2 for desirable (more petals per flower, greater flowering, shorter plants), undesirable (less flowering, smaller flowers), and neutral (narrower leaves, taller plants) traits. Open-pollinated (OP) seed was collected and germinated and the seedlings from somaclones that differed significantly from E2 and H2 were evaluated. These maternally selected seedlings were overwintered then planted in the field in May 2004. Most traits that differentiated somaclones from E2 and H2 did not persist in the OP seedling population; however variation that was likely introduced through outcrossing resulted in desirable phenotypes with potential for new cultivar development.

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Dirk R. Vuylsteke, Rony L. Swennen, and Edmond A. De Langhe

Four types of morphologically distinct somaclonal variants were identified in a population of False Horn plantain (Musa spp., AAB group) plants produced by in vitro shoot-tip culture. Field performance of these variants was compared with true-to-type plantain to evaluate their horticultural traits. Significant variation was observed for plant and fruit maturity, leaf size, yield and its components, but not for leaf number, plant height, or suckering. Three of the four somaclonal variants were horticulturally inferior to the original clone from which they were derived. Yields of these variants were very poor due to inflorescence degeneration or abnormal foliage. Only the `French reversion' variant, which resembled an existing cultivar, outyielded the true-to-type clone. However, its fruit weight and size were lower. Somaclonal variation through micropropagation is of limited use in plantain improvement as it mostly mimics naturally occurring variation along with the observed poor horticultural performance of somaclonal variants.

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Dae-Geun Oh and Edward C. Tigchelaar

The tangerine-virescent (tv) mutation was reported as a single gene somaclonal variant from tissue culture (Evans and Sharp 1963). A replicated field trial was conducted to characterize variation and stability in the phenotype of this tv somaclone and to compare it with the inbred parent from which it was reportedly derived.

Heritability and stability of the tv somaclonal variant was measured by comparing R3 end R4 lines of sexual progenies of the original tv variant and with its sexually derived inbred parent UC82B. Several additional variants were observed in these tv lines, including fruit shape, days to first flower, fruit weight, yield, plant type, and fertility. Eight sterile or semi-sterile plants were discovered in 6 of 39 R4 lines. Our results suggest that multiple genetic changes have occurred in the tv somaclonal variant and while the original tv mutant is stably inherited, additional genetic abnormalities occur following sexual reproduction.

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Stan C. Hokanson, Kelvin G. Grant, Elizabeth L. Ogden, and Lisa J. Rowland

Commercial strawberry plantings in the mid-Atlantic region are often quickly infected with one or more aphid-transmitted viruses, resulting in the loss of plant vigor, stunting, lowered yields, etc. To produce virus-free plant material for the strawberry industry and for cultivar development programs, heat therapy and/or meristem tip culture protocols are generally employed. One of the problems associated with meristem culturing is the potential for somaclonal mutations to occur in the meristem or surrounding proliferating tissue. As a result, distinct “bud lines” displaying functionally insignificant to distressingly high levels of phenotypic variation can arise from individual meristems. It would be desirable to differentiate these off-types by genetic fingerprinting to maintain trueness-to-type. Randomly amplified polymorphic DNA (RAPD) markers were evaluated for the potential to differentiate six pairs of strawberry bud lines that exhibit slight to fairly extreme levels of phenotypic variation. Reproducible RAPD marker profiles were generated using 10 primers in amplification reactions with genomic DNA obtained from multiple extractions. While five of the bud line pairs remained indistinguishable, three primers distinguished two variants of the Mohawk cultivar that are now in existence in the strawberry industry. Results suggest that typical somaclonal variation produced in the meristem culture process is of a magnitude that is not readily detectable with the RAPD protocol. The two Mohawk lines were probably produced by a higher magnitude mutation event than generally occurs or a cultivar mix-up.