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Xiaoming Wang, Yongxin Li, Huijie Zeng, Neng Cai, Zhongquan Qiao, Xiangying Wang, and Jianjun Chen

) regenerated plantlets of W. florida ‘Bristol Ruby’ using protoplasts as explants. The MS medium was also used for shoot culture of W. florida ‘Red Prince’, but vitrification was a problem ( Wang et al., 2000 ). Additionally, callus was induced from ‘Red

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Eric L. Zeldin, David D. Ellis, and Brent H. McCown

Taxol, a promising anticancer drug, is limited by inadequate supply. The production of taxol and related compounds (taxanes) by Taxus tissue cultures has been reported, yet sustained production has not been demonstrated. One theory is that cell differentiation and/or tissue organization is required to sequester taxol and avoid autotoxicity. To investigate this, T. cuspidata shoot cultures were established and the taxane content of various culture stages compared to that of field needles. HPLC analysis identified two peaks which comigrated and had UV spectra identical to taxol and 10-deacetyl taxol. The levels of 10-deacetyl taxol were similar in all samples. Cultured shoots contained much less taxol than field needles, and the level of a third peak which migrates closely to taxol was inversely related to that of taxol. Taxol content was restored in the first flush out of culture. Shoot cultures of T. brevifolia, T. x media, and T. canadensis have also been analyzed. In addition to shoot cultures, nodule cultures, a biological unit that may be suitable for production of taxanes in plant bioreactors, have been initiated and characterized.

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Michael Marcotrigiano, Thomas H. Boyle, Pamela A. Morgan, and Karen L. Ambach

Nuclear-controlled leaf variegation was studied among Coleus × hybridus Voss (formerly C. blumei Benth.) cultivars propagated by seed and as shoot cultures on Murashige and Skoog (MS) medium + 1 to 3 mg BA/liter. Cultivars tested possessed pattern chlorophyll variegation and either pattern or nonpattern anthocyanin variegation. The gene controlling an albino midrib region appears to be fairly stable, with only 2% of the micropropagated plantlets having a solid-green leaf characteristic, a characteristic that was always inherited following selfing. Pattern anthocyanin variegation (PAV) was fairly stable, while nonpattern anthocyanin variegation (NAV) was very unstable. In addition, variants from pattern-variegated phenotypes produced offspring identical to their parent following selfing. In contrast, variants of nonpattern cultivars, when selfed, yielded offspring identical to the original cultivar, identical to the variant, or novel phenotypes. When variants were returned to culture, those derived from cultivars with PAV were more stable than those derived from nonpattern cultivars. In Coleus, micropropagation may induce epigenetic and/or heritable changes in leaf variegation. Cultivars with NAV are less stable than cultivars with PAV. Chemical names used; N-(phenylmethyl)-lH-purine-6-amine [benzyladenine (BA)].

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P.A. O`Connor and S.S. Korban

Established shoot cultures of three apple genotypes, `Dayton', `McIntosh', and `Golden Delicious' were subcultured into culture tubes containing a modified MS medium and maintained in a dark chamber at 1.0±0.5°C for periods of 3, 6, 9, and 12 months. Following each cold storage period, culture tubes of each of the three genotypes were transferred to a growth room and maintained under 16 h of light (60 uEs-1m-2) and 21°C. The overall morphological condition of each shoot was then recorded. After 4 weeks of growth, both number and length (in cm) of proliferating shoots were recorded. In general, shoots subjected to 3 or 6 months of cold storage remained green however most cultures did not initiate any new shoots. Cultures subjected to 9 or 12 months of cold treatment were etiolated however new axillary shoots were observed. The proliferation rate after 4 weeks of growth under standard growth conditions were variable among the different genotypes. The implications of using long term cold storage of apple shoot cultures will be discussed.

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P.A. O`Connor and S.S. Korban

Established shoot cultures of three apple genotypes, `Dayton', `McIntosh', and `Golden Delicious' were subcultured into culture tubes containing a modified MS medium and maintained in a dark chamber at 1.0±0.5°C for periods of 3, 6, 9, and 12 months. Following each cold storage period, culture tubes of each of the three genotypes were transferred to a growth room and maintained under 16 h of light (60 uEs-1m-2) and 21°C. The overall morphological condition of each shoot was then recorded. After 4 weeks of growth, both number and length (in cm) of proliferating shoots were recorded. In general, shoots subjected to 3 or 6 months of cold storage remained green however most cultures did not initiate any new shoots. Cultures subjected to 9 or 12 months of cold treatment were etiolated however new axillary shoots were observed. The proliferation rate after 4 weeks of growth under standard growth conditions were variable among the different genotypes. The implications of using long term cold storage of apple shoot cultures will be discussed.

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James S. Busse, Senay Ozgen, and Jiwan P. Palta

typified by the browning and death of the shoot tip, the loss of apical dominance, and axillary branch development in an in vitro shoot culture system ( McCown and Sellmer, 1987 ; Sha et al., 1985 ). Transpiration is limited during in vitro culture by high

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Areej A. Alosaimi, Robert R. Tripepi, and Stephen L. Love

38 μmol·m −2 ·s −1 photosynthetic photon flux ( PPF ). After 4 weeks, shoot cultures were subcultured and placed on fresh MS medium or WPM two times before axillary shoot proliferation was started. Because stem explants grew best on the MS salt

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Basdeo Bhagwat and W. David Lane

The insecticides acephate, dichlorvos, and imidacloprid were assayed, using in vitro shoot cultures of apple (Malus×domestica Borkh.), to determine their phytotoxicity at several concentrations and their effectiveness for eradication of the Western Flower Thrip (Frankliniella occidentalis, Pergande) from infested apple shoot cultures. Commercial formulations of acephate (Orthene), dichlorvos (VaportapeII), and imidacloprid (Admire) and a technical grade of imidacloprid were used in the experiments. For acephate and imidacloprid, concentrations of 1 to 80 mg·L-1 a.i. in shoot culture medium were used, while for dichlorvos, a fumigant, particles of the formulated product containing concentrations of 0.7 to 6.4 mg a.i. were suspended in the head space of the 500-mL glass culture jar. Acephate, dichlorvos, and the technical grade of imidacloprid did not cause phytotoxicity and growth of shoot cultures was unaffected at all treatment concentrations tested after a 6-week treatment period. Imidacloprid (20 to 80 mg·L-1 of the commercial formulation) caused chlorosis at the end of the 6-week treatment period. None of the treatments tested resulted in the death of shoots. Thrips were eradicated by acephate or imidacloprid treatments of 5 mg·L-1 and by dichlorvos treatment of 0.7 mg per 500-mL culture jar. Shoot cultures grew normally after the treatment period. Chemical names used: O,S-dimethyl acetylphosphoramidothioate (acephate), 2,2- dichlorovinyl dimethyl phosphate(dichlorvos),1-(6-chloro-3-pyridylmethyl)-N-nitroimidazolidin-2-ylideneamine (imidacloprid).

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Arthur M. Richwine, Jimmy L. Tipton, and Gary A. Thompson

Shoot cultures of Aloe, Gasteria, and Haworthia species were initiated directly from immature inflorescences. Explants placed on a modified MS medium containing 5.4 mm zeatin riboside initiated shoots within 8 to 12 weeks. Long-term shoot cultures were established and maintained on media containing either 5.4 μm zeatin riboside or 4 μm BA. Shoots easily rooted in vitro, and rooted plantlets were esablished in soil. Chemical names used: N-(phenylmethyl)-1H-purin-6-amine (BA); 6-[4-hydroxy-3-methyl-but-2-enylamino]purine riboside (zeatin riboside).

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Nan Wang and Barbara M. Reed

Roots of greenhouse-grown mint plants and in-vitro-grown shoot cultures were inoculated with Verticillium dahliae Kleb. conidial suspensions to study wilt symptom development and detection and elimination of the fungus. There were significant differences in the symptom expression between control and infected shoot cultures at all conidia concentrations for the four mints tested. Disease-symptom ratings were proportional to the V. dahliae inoculum density. Infected shoot cultures were stunted when inoculated with ≥ 103 conidia/mL. Verticillium dahliae was re-isolated from infected shoot cultures at all levels of inoculum, but not from any control cultures. Verticillium infections were easily detected by plating mint stems on potato dextrose agar. Shoot tips (0.5 to 15 mm) from infected in-vitro- and greenhouse-grown plants were isolated and screened for fungus. The most effective shoot length for fungus elimination was 3-5 mm. Shoot tips isolated from in vitro spearmint cultivars infected at 102 and 103 conidia/mL were 100% Verticillium free, but only 42% of `Black Mitcham' and 54% of `Todd's Mitcham' peppermints were free of the disease. Shoot tips from infected greenhouse plants produced Verticillium-free cultures from 79% of `Black Mitcham' and 90% of `Todd's Mitcham' plants. These results indicate the utility of testing for Verticillium and the safety of micropropagated mint shoots for certified planting stock programs.