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Lauren E. Deans, Irene E. Palmer, Darren H. Touchell, and Thomas G. Ranney

), and H. serrata ‘MAK20’ ( C ) plants during their third ex vitro growing season. A comparison of floral characteristics showed that induced tetraploidy caused a significant decrease in inflorescence number for all cultivars ( P < 0.05), with an

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John L. Edson, David L. Wenny, and Annette Leege-Brusven

In vitro—derived microshoots of antelope bitterbrush, incubated for 1 month in media supplemented with 0.44 μm BA, grew 0.8 and 1.1 cm longer in woody plant medium (WPM) compared to full-strength and half-strength Murashige and Skoog (MS) media, respectively. Explants cultured in WPM supplemented with 0.44 μm BA and 0.54 μm NAA produced a mean of five axillary shoots per explant. Explants dipped in 0.1% IBA or 0.1% NAA rooted best in 0.1% IBA with 89% success ex vitro vs. 60% success in vitro. Survival of acclimatized plantlets rooted ex vitro was 95%, while 50% survived when rooted in vitro. After 1 year of greenhouse growth, 98% of plantlets survived and flowered. Chemical names used: benzyladenine (BA), 3-indolebutyric acid (IBA), 1-naphthaleneacetic acid (NAA).

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Yulan Xiao and Toyoki Kozai

A photoautotrophic or sugar-free medium micropropagation system (PAM) using five large culture vessels (volume = 120 L each) with a forced ventilation unit for supplying CO2-enriched air was developed and applied to commercial production of calla lily (Zantedeschia elliottiana) and china fir (Cunninghamia lanceolata) plantlets. The culture period of calla lily plantlets in the PAM was reduced by 50%, compared with that in a conventional, photomixotrophic micropropagation system (PMM) using small vessels each containing a sugar-containing medium. Percent survival ex vitro of calla lily plantlets from the PAM was 95%, while that from the PMM was 60%. The production cost of calla lily in the PAM was reduced by about 40%, compared with that in the PMM, and the initial investment per plantlet for the PAM was ≈10% lower than that for the PMM. The sales price of ex vitro acclimatized calla lily plantlet was increased by 25% due to its higher quality, compared with plantlets produced in the PMM.

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Ribo Deng and Danielle J. Donnelly

Micropropagated `Festival' red raspberry (Rubus idaeus L.) shoots were rooted in specially constructed plexiglass chambers in ambient (340 ± 20 ppm) or enriched (1500 ±50 ppm) CO2 conditions on a medium containing 0, 10, 20, or 30 g sucrose/liter. Plantlet growth and leaf 14CO2 fixation rates were evaluated before and 4 weeks after ex vitro transplantation. In vitro CO2 enrichment promoted in vitro hardening; it increased root count and length, plantlet fresh weight, and photosynthetic capacity but did not affect other variables such as plantlet height, dry weight, or leaf count and area. No residual effects of in vitro CO2 enrichment were observed on 4-week-old transplants. Sucrose in the medium promoted plantlet growth but depressed photosynthesis and reduced in vitro hardening. Photoautotrophic plantlets were obtained on sucrose-free rooting medium under ambient and enriched CO2 conditions and they performed better ex vitro than mixotrophi plantlets grown with sucrose. Root hairs were more abundant and longer on root tips of photoautotrophic plantlets than on mixotrophic plantlets. The maximum CO2 uptake rate of plantlet leaves was 52% that of greenhouse control plant leaves. This did not change in the persistent leaves up to 4 weeks after ex vitro transplantation. The photosynthetic ability of persistent and new leaves of 4-week-old ex vitro transplants related neither to in vitro CO2 nor medium sucrose concentration. Consecutive new leaves of transplants took up more CO2 than persistent leaves. The third new leaf of transplants had photosynthetic rates up to 90% that of greenhouse control plant leaves. These results indicate that in vitro CO2 enrichment was beneficial to in vitro hardening and that sucrose may be reduced substantially or eliminated from red raspberry rooting medium when CO2 enrichment is used.

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Suzanne M. Dethier Rogers and Sharon Banister

A micropropagation system was developed for micropropagation and rooting of Notholaena spp. Shoot cultures were initiated from mature sori on leaflets of the Notholaena cultivar Sun-Tuff and cultured on gelled Murashige and Skoog (MS) medium without hormones. Rooting and plant growth were evaluated on gelled MS, vermiculite moistened with liquid MS, or vermiculite moistened with water. Vermiculite wetted with MS was superior in promoting frond and root development. High humidity was not needed to acclimatize the plants ex vitro.

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Andres A. Estrada-Luna, Fred T. Davies Jr., and Jonathan N. Egilla

The role of mycorrhiza fungi during acclimatization and post-acclimatization of micropropagated chile ancho plantlets was characterized through physiological and plantlet development changes. Regardless of mycorrhizal colonization, the pepper plantlets had initially low photosynthetic rates and poor growth following transplanting ex vitro. During the first days of acclimatization, water deficits occurred as evidenced by drastic reductions in relative water content. Consequently, transpiration rates and stomatal conductance (gs) declined, confirming that in vitro formed stomata were functional, thus avoiding excessive leaf dehydration and plant death. Mycorrhiza had a positive effect on gas exchange as early as day 7 and 8, as indicated by increasing photosynthesis (A) and gs. Mycorrhizal plantlets had reduced levels of abscisic acid (ABA) during peak stress (6 days after transplanting ex vitro), which corresponded with subsequent increases in gs and A. During acclimatization, A increased in both non-colonized and colonized plantlets, with greater rates observed in mycorrhizal plantlets. During post-acclimatization, mycorrhiza colonized 45% of the roots of pepper plantlets and enhanced plant growth by increasing leaf area, leaf dry mass, and fruit number. Mycorrhiza also enhanced total leaf chlorophyll content, A, and nutrient uptake of pepper plantlets, particularly N, P, and K. Early mycorrhizal colonization produced important benefits, which helped ex vitro transplanted plantlets recover during acclimatization and enhance physiological performance and growth during post-acclimatization.

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Jeffrey Adelberg, Kazuhiro Fujiwara, Chalermpol Kirdmanee, and Toyoki Kozai

Two triploid clones of melon from the same tetraploid parent were grown in vitro with and without sugar, rooted without sugar in media both in a laboratory controlled environment chamber (in vitro) and a greenhouse acclimatization unit (ex vitro), and compared for subsequent nursery growth in the greenhouse unit. The clone `(L-14 c B) × L-14' produced more shoots in both photomixotrophic (with sucrose) or photoautotrophic (sugar-free) conditions. Both genotypes were equally likely to root in sugar-free media, and `(L-14 × B) × L-14' rooted as well from either photoautotrophic and photomixotrophic shoots but `(L-14 × B) × Mainstream' rooted less frequently from photoautotrophic shoots. Seventy-six percent (76%) of the shoots were able to root photoautotrophically in vitro, whereas 47% of the ex vitro shoots were rooted. About 85% of plantlets from all treatments survived after transfer to the nursery. After growth in the greenhouse nursery, the sizes of plants (fresh and dry weight, leaf area) were the same for either clone, from either photoautotrophic or photomixotrophic shoots. Also, after growth in the nursery, plantlets that had been rooted in vitro were larger than those rooted ex vitro. Photoautotrophic rooting demonstrates a concept for integrating micropropagation and plug-type vegetable transplant production.

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Mehmet Nuri Nas and Paul E. Read

Microshoots of four hazelnut genotypes grown in vitro on Nas and Read medium (NRM) containing various combinations of CuSO4 • 5H2O and myo-inositol were successfully rooted and acclimatized ex vitro without any need of in vitro hardening treatments. Dipping of shoot bases in 1000 ppm indole-3-butyric acid (IBA) solution for 5 or 10 seconds followed by placement of shoots in plant growth regulator free NRM gave rise to formation of roots as early as 8 days. Shoots treated for 5 and 10 seconds rooted similarly, and depending on genotype, 88% to 98% rooting was observed within 15 days after treatment with IBA. Ex vitro survival of shoots three months after in vitro-root induction was 73% when shoots were treated with IBA for 5 seconds and 66% when shoots were treated for 10 seconds. The highest ex vitro survival rate (97%) 3 months after root induction was observed when shoots were treated with IBA solution for 10 seconds, and then cultured directly in peat pellets. Shoots developed good roots, and grew up to 70 cm in height 3 months after root induction. The potential use of rooting and acclimatization protocol for commercial micropropagation of hazelnut is presented.

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Martine Korban and Danielle J. Donnelly

Mechanically induced stress (shaking stress) applied during shoot multiplication (Stage II) or rooting (Stage III) of micropropagated `Queen Elizabeth' rose was evaluated to determine its effects on in vitro hardening. Shaking during Stage II did not alter the growth responses of the shoots before transfer to Stage III. Shaking during Stage III, at 150 rpm for 15 min daily for 2 weeks, only caused a reduction in leaf dry weights before transfer to soil. Automated shaking stress during Stages II or III did not apparently promote hardening of cultured plants or improve their ex vitro performance.

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Ribo Deng and Danielle Donnelly

Labeled (`“C) compounds were recovered from tissue disks taken from 14CO2-fed leaves of l-year-old greenhouse-grown plants and l-month-old ex vitro transplants of red raspberry (Rubus idaeus L.) by hot (boiling in 80% ethanol immediately after 14C exposure), delayed-hot (boiling in 80% ethanol after a 2- to 3-day ethanol soak), and room-temperature (RT) (2-to 3-day soak in 80% ethanol) extraction methods. The RT extraction method was simple but as effective for extracting 14C-labeled compounds from red raspberry leaf tissues as hot and delayed-hot extraction methods.