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V.R. Bommineni, H. Mathews, S.B. Samuel, M. Kramer, and D.R. Wagner

Improved in vitro clonal propagation methods are valuable tools for nurseries and growers, and are essential for manipulation and improvement of tree fruit germplasm using the tools and techniques of biotechnology. We have developed a rapid shoot multiplication procedure for clonal propagation of apple, Malus ×domestica cv. Gale Gala and pear, Pyrus communis L. cv. Bartlett. Rapid clonal multiplication was achieved after the following series of steps: pre-conditioning of micropropagated shoots, sectioning pre-treated stems into thin slices, placing slices onto shoot induction medium and incubating directly under cool-white fluorescent lights or after a brief dark incubation. Multiple induction of shoots recovered from stem slice explants within three weeks of culture. A maximum of 37% of cultured apple stem slices, and 97% of pear stem slices, showed induction of shoots. More shoots were recovered on phytagel solidified shoot induction medium than on agar. Cultured stem slices of both apple and pear showed maximum recovery of shoots from shoot induction medium supplemented with thidiazuron (TDZ) compared to medium supplemented with BAP and kinetin. Under ideal conditions, pear stems generated four times the shoots as the same quantity or length of apple shoots. Micropropagated shoots were rooted and transferred to the greenhouse and field nursery for further evaluation. Chemical names used: N-phenyl-N′-1,2,3-thidiazol-5-ylurea (thidiazuron or TDZ); 6-benzylaminopurine (BAP).

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Nancy Santana-Buzzy, Adriana Canto-Flick, Felipe Barahona-Pérez, María del Carmen Montalvo-Peniche, Patricia Yolanda Zapata-Castillo, Anabel Solís-Ruiz, Amílcar Zaldívar-Collí, Omar Gutiérrez-Alonso, and María de Lourdes Miranda-Ham

To induce multiple shoots from habanero pepper (Capsicum chinense Jacq.), nodes and stem segments were cultivated in MS medium supplemented with varying concentrations of kinetin, benzyladenine, and thidiazuron. The effect of the age of the explant in the medium on shoot formation and their latter development into plants was assessed. Ethylene concentration was measured along the experiments. Thidiazuron was the key growth regulator in the process, which at 3.4 μm induced seven to eight shoots that developed into healthy plants per explant. Plantlets in nonventilated vessels, where ethylene concentration was 0.25 ± 0.1102 μL·L–1, showed early defoliation and the formation of calli on the leaves and stems.

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Hak Tae Lim and E.J. Park

The regeneration medium supplemented with 2.0 mg/L BAP and 0.1 mg/L IAA allowed high efficient shoot regeneration from leaf discs and petioles of Cichorium intybus L. var. sativus. Multiple shoots ranged from 10 to 14 per explant were observed only 10 to 15 days after the initial culture. Reduced nitrogen and sucrose levels influenced on shoot regeneration frequency and growth rates. Especially, in C. Intybus L. var. sativus cv. Cesare explants cultured in the medium containing 50 mg/L MS macroelement and 1.5% sucrose displayed high regeneration frequency of 100%.

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Yaseen Mohamed-Yaseen, Raymond J. Schnell, Robert J. Knight, and T. L. Davenport

Guava (Psidium guajava L.) is an exceptional source of vitamin. C. It is also considered to be the most important cultivated species of the Myrtel family. Shoot tip and stem node were taken from seedling germinated in Murashige and Skoog medium (MS) and cultured in the same medium supplemented with 1-3mg/l benzylaminopurine (BA) and 0.1mg/l naphthaleneacetic acid (NAA) or 0.2-2mg/l thidiazuron (TDZ) and 0.1mg/l NAA. Multiple shoots (4-6) were obtained in 4-5 weeks from culture in 1-2mg/l BA and 0.1mg/l NAA, while TDZ caused abnormal shoot growth. Shoots were rooted successfully with 100% frequency in MS medium containing 2mg/l indolebutyric acid and further elongation of shoots was achieved in MS medium, supplemented with lg/l activated charcoal. Regenerated plantlets were successfully established in soil.

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Kee Yoeup Paek and Toyoki Kozai

We report the results of serial studies aimed at clarifying several factors affecting organogenesis in rhizome culture of temperate Cymbidium species and their hybrids. The growth patterns and regeneration ability of rhizomes derived from asymbiotic seed or shoot tip culture vary according to media composition, kinds and concentrations of plant growth regulators, culture conditions, and species and varieties. N6-benzyladenine was the best cytokinin for inducing shoot formation, for switching rhizome tissues into protocorm-like bodies, and for directly forming multiple shoots from branched rhizomes. Activated charcoal appeared to be necessary for producing healthy plantlets and for stimulating shoot growth at levels of 0.1% to 0.3% but concomitantly decreased rhizome growth. Sucrose at 5% was the most effective concentration for shoot induction from rhizomes. The above results support the conclusion that organogenic pathways between tropical, subtropical, and temperate Cymbidium species may be controlled by the genetic backgrounds of the species or cultivars.

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E.E. Chesick, D.E. Bilderback, and G.M. Blake

Vegetative long-shoot buds, greenwood stems, and immature needles of 20-year-old western larch (Larix occidentalis Nutt.) were cultured to induce multiple bud formation. Explants were collected year-round and cultured on a modified Schenk and Hildebrandt (SH) medium containing 6-benzyladenine (BA) at 0, 1, 5, 10, 50, or 100 μm. Multiple buds were produced on buds and stems with terminal meristems, but not on needles or stem sections. The induction of de novo buds and development of axillary buds required BA at 1 to 10 μm; higher concentrations of BA were less effective. More explants formed multiple buds on SH than on modified Murashige and Skoog (MS) media. Multiple buds formed on more buds and stems excised during the growing season than from dormant buds. Buds cultured on media containing gibberellin died within 6 weeks; auxin caused bud elongation but no multiple buds formed. Chemical names used: N-[(trichloromethyl)thio]-4-cyclohexene-1,2-dicarboximide (captan); 6-benzyladenine (BA); 1H-indole-3-butyric acid (IBA); 1H-indole-3-acetic acid (IAA); gibberellin (GA4+7).

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Anne E. Sama and Harrison G. Hughes

Shoot tips, approximately 3-5mm, were isolated from corms of young greenhouse-grown plants of cocoyam, cultivar South Dade White. After preliminary evaluations, the initiation media evaluated were B5 basal salts supplemented with 0.05 μM NAA with 5μM BAP, 20μM BAP or 2μM TDZ. The above media were in the form of liquid medium in flasks on a rotary shaker, liquid medium with filter paper bridges, stationary liquid medium without filter paper and solidified medium with 0.4% agar. TDZ stimulated greater growth with multiple shoot formation. Liquid media either in the shaker or stationary form were more effective in terms of growth. Shoots were subsequently evaluated for multiplication with 1μM TDZ and 5μM BAP with 0.05μM NAA producing greater shoot numbers. Over 30 plants have subsequently been rooted and acclimatized under mist or humidity tent.

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M.K. Ehlenfeldt, A.W. Stretch, and V. Brewster

The resistance of 48 highbush blueberry cultivars and selections to the blight phase of mummy berry disease, incited by the fungus Monilinia vaccinii-corymbosi (Reade) Honey, was examined in relation to percent Vaccinium angustifolium Ait. ancestry, season of fruit maturity, and shoot growth during the primary infection phase. Correlations of percent blighting with percent V. angustifolium ancestry were significant across 3 years, but correlations with fruit maturity were significant in only 2 of 3 years. Correlations of percent blighting with early shoot growth were significant in both years measured, with r values of 0.54 in 1994, 0.83 in 1995, and 0.83 across years. A multiple regression found only shoot growth highly significant for susceptibility and rendered V. angustifolium ancestry and season of fruit maturity nonsignificant. Resistant cultivars exhibiting early shoot elongation suggest that resistance can be either biochemically or escape based.

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Michael E. Kane and Charles Lane

Many wetland plant species used for aquascaping and wetland revegetation projects are collected from donor wetland sites for planting elsewhere. Increased demand for wetland plants has lead to over-collection and subsequent environmental damage to these donor sites. Micropropagation provides an ecologically sound alternative to field collection and allows for production of under utilized wetland species and genotypes that are either slow-growing or difficult to propagate using conventional methods. Sagittaria latifolia Willd. (Duck-potato), a rhizomatous herbaceous wetland species, was established in vitro from surface-sterilized lateral and terminal rhizome shoot-tips cultured in liquid basal medium consisting of half-strength Murashige and Skoog mineral salts, 0.56 mM myo-inositol and 1.2 μM thiamine supplemented with 87.6 mM sucrose. Prior to multiplication, responsive Stage I cultures were indexed for cultivable bacteria and fungi. Shoot multiplication occurred in vitro through formation of multiple node rhizomes bearing terminal shoots. Duck-potato exhibited a high sensitivity to relatively low benzyladenine (BA) levels. Maximum rhizome and shoot production occurred from single shoot explants initially cultured on agar-solidified BM supplemented with 4.0 μM BA for 28 days. However, repeated subculture on BM supplemented with greater than 2.5 μM BA resulted in increased mortality, reduction in multiplication rate, or production of dormant corms. Consistent shoot multiplication (four to five shoots/explant) was possible in the presence of 1.5 μM BA. Maximum (100%) acclimatization and rooting was attained by direct sticking of Stage II microcuttings in soilless growing medium contained in 38 cell plugs. Production of salable plants bearing multiple rhizomes was possible within 6 weeks post-transplant. Preliminary observations indicate that corm formation in Sagittaria latifolia may be mediated by photoperiod.

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John M. Ruter

A study was conducted with Prunus × incamp `Okame' to evaluate the effects of a pot-in-pot production system compared to a conventional above-ground system and cyclic irrigation on plant growth and water loss. Plants were grown in #7 (26-L) containers with a 8:1 pinebark:sand (v/v) substrate. Cyclic irrigation provided the same total volume of water, but was applied one, three, or four times per day. Final plant height and stem diameter, shoot and root dry weight, total biomass, and root:shoot ratio were all increased for plants grown pot-in-pot compared to above-ground. Multiple irrigation cycles increased stem diameter, shoot dry weight, and total biomass, compared to a single irrigation application. Multiple irrigation cycles decreased the root:shoot ratio. Evapotranspiration was influenced by production system, irrigation, and date. Amount of water lost as leachate was influenced by irrigation and date. Cyclic irrigation resulted in a two-fold decrease in leachate volume. Soluble salts and nitrate-nitrogen in the leachate were influenced by an interaction between production system, irrigation, and date.