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Kh. A. Okasha and M. E. Ragab

177 POSTER SESSION 25 (Abstr. 870-901) Cell and Tissue Culture/Propagation

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Vyacheslav Gurevich, Uri Lavi, and Yuval Cohen

Date palm (Phoenix dactylifera L.) is a major tree crop in arid regions of the Middle East and North Africa, having an important impact on the economy of many countries in these regions. Date palms are traditionally propagated through offshoots. The development of propagation methods through tissue culture resulted in massive expansion of date palm plantations. While most trees generated from tissue culture are normal and true-to-type, several typical abnormal phenotypes are detected. The present study applies amplification fragment length polymorphism (AFLP) analysis to characterize the genetic variation of two elite date cultivars, `Barhee' and `Medjool', as well as male clones, propagated from offshoots and through tissue culture. The two cultivars have very distinct AFLP band patterns. Most offshoots, as well as the tissue culture-propagated plants, have very similar band patterns, demonstrating a low level of genetic variation. However, a significant level of genetic variation was detected among `Medjool' plants generated from tissue culture. Several phenotypically abnormal trees were characterized by unique and different AFLP band patterns. The male clones are characterized by a high level of polymorphic bands. Genetic variation was also detected between various tissues of variegated `Medjool' trees propagated from tissue culture. The significance of these results, regarding the mechanism of the phenomenon and its relevance to agricultural practice, is discussed.

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Renee DeVries-Paterson, Christine Stephens, and Thomas Evans

Commercial asparagus (Asparagus officinalis L.) is currently planted from seed but there is a growing interest in the use of tissue culture clones. The worldwide occurrence of asparagus virus I (AV-I) and asparagus virus II (AV-II) in asparagus production areas has led to an investigation of the effect of these viruses singly and in combination on the propagation of asparagus via tissue culture. Bud explants from field-grown, virus-infected asparagus plants were cultured in-vitro to induce shoots and roots. Explants derived from singly or doubly-infected plants were slow to develop roots and often died in culture. The four virus groups were ranked for the explants' capacity to produce roots and shoots: virus-free > AV-II > AV-I> AV-I and AV-II. Plants derived from explants of AV-II-infected plants exhibited a mild weight reduction after three months in the greenhouse. Greater reductions were associated with AV-I and double infections when compared to healthy controls.

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Scott J. Nissen and Ellen G. Sutter

The relative stabilities of IAA and IBA under various tissue culture procedures were determined. IBA was significantly more stable than IAA to autoclaving. IBA was also found to be more stable than IAA in liquid Murashige and Skoog medium (MS) under growth chamber conditions. The stabilities of IBA and IAA were similar in agar-solidified MS. Light provided by cool-white fluorescent bulbs promoted degradation of IAA and IBA in both liquid and agar media. Activated charcoal in concentrations as high as 5% was found to adsorb more than 97% of IAA and IBA in liquid MS. These results have important implications for the preparation, storage, and handling of IBA and IAA in plant tissue culture. Chemical names used: indole-3-acetic acid (IAA); indole-3-butyric acid (IBA).

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Rita M. Moraes-Cerdeira, Jeffrey V. Krans, James D. McChesney, Ana M.S. Pereira, and Suzelei C. Franca

Cotton fibers were tested as a substitute for agar in tissue culture. The cost of agar has prompted us to search for an alternative more economical medium support. Effectiveness as a medium support was evaluated in terms of callus maintenance and shoot organogenesis using Artemisia, Agrostis, and Taxus. Taxus and Agrostis calli cultivated on liquid media with cotton fiber as medium support (25 ml of medium per gram of cotton) grew better than calli on agar (0.8% w/v). There were no significant differences in shoot organogenesis of Artemisia and Agrostis grown in 25 ml of medium per gram of cotton from those grown in agar medium.

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Chi Won Lee, Chun-Ho Pak, and Harrison G. Hughes

Stevia (Stevia rebaudiana Bert.) leaves produce stevioside and rebaudioside that can be used as a natural source of low-calorie sweetener which is heat-stable. Because of low fertility, this plant is often vegetatively propagated for field production. This study was conducted to optimize tissue culture procedures for propagating selected clones and explore the feasibility of producing the sweetener compounds by callus cultures. Shoot proliferation was best in Murashige and Skoog (MS) medium supplemented with 0.1 mg/l naphthaleneacetic acid (NAA) Plus 10 mg/l kinetin. Kinetin as a cytokinin source was better than benzyladenine (BA) especially when NAA was present. Callus production fronm leaf disc cultures was most prolific when a combination of 0.1 mg/l NAA and 3 mg/l BA was used in MS medium. The relative sweetener contents of callus cultures are currently being-analyzed.

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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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Michael E. Compton and Richard E. Veilleux

Genetic recombination rates of hybrid plants regenerated from three tissue culture. systems were compared by backcrossing regenerated plants with mutant parents and comparing the observed crossover frequencies with those expected based on control plants raised from seed. Increased recombination rates and map distances were observed among plants from micropropagated shoot tips (4.5%-5.9%), cotyledon calli (3.7%-8.5%), and thin cell layers (2.8%-6.5%) between the sunny (sy) and baby leaf syndrome (bls) markers which flank the centromere on chromosome 3. Conversely, a decrease in map distance was observed between bls and the solanifolia (sf) locus which is more distal to the centromere on the same arm of chromosome 3 as bls. Increased map distance among plants regenerated from micropropagated shoot tips, cotyledon calli, and thin cell layers was also observed between white virescence (wv) and anthocyanin reduced (are) loci on chromosome 2.

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Kenneth W. Mudge and Chin-Chang Chu

In vitro asymbiotic seed germination, subculture, and outplanting of orchids is presented as a laboratory exercise suitable for students of plant propagation or tissue culture. Dendrobium antennatum (Lindley), Phalaenopsis (Blume) white hybrid, or both, are used in this exercise because they flower predictably in the greenhouse, are reliable for seed production, and germinate and grow rapidly in vitro. The exercises can be used to instruct students in the skills involved in orchid seed sterilization, sowing, and culture, as well as instruct students in the unique features of orchid reproductive biology and symbiosis. A schedule is suggested for stock plant flower pollination, capsule harvest, seed sowing, and seedling subculture so that the necessary plant material is available for students to sow, subculture, and outplant seedlings during a single laboratory session.

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Denise M. Seliskar

Several salt marsh functions were found to be directed by the genotype of the vascular plant. Tissue culture regenerants of Distichlis spicata and Sporobolus virginicus, along with plants from natural populations of these species, were compared in a common garden study. Significant differences among genotypes were found in several characteristics of importance in the functioning of the salt marsh food web. Specifically, potential detritus production, belowground organic matter production, and canopy structure were affected. Selections from five morphologically distinct populations of Spartina patens, including one tissue culture regenerant, also maintained differences in a common marsh setting. In a newly created salt marsh near Lewes, Dela., three populations of short-form Spartina alterniflora from different latitudes (Massachusetts, Delaware, and Georgia) were planted. After 5 years in the new site, the plants maintained distinct morphologies characteristic of plants at their site of origin; e.g., aboveground biomass, canopy structural characteristics, and root and rhizome biomass, composition, and distribution. The magnitude of marsh functional processes associated with the latitude of the population source persisted in the created wetland. The edaphic algal community, the activity of the aerial microbial decomposers, and the edaphic community respiration were significantly influenced by vascular plant genotype in the created marsh, and they reflected the values characteristic of these functions at the latitude of origin of the vascular plants. Indications are that higher trophic levels may also be affected. In creating new wetlands, this plant genetic variation can be used to accentuate different functions, thus optimizing wetland values on the specific site and at the landscape ecology scale.