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John E. Preece and Carl A. Huetteman

This exercise was developed for a plant propagation course to demonstrate, in a short time, the four stages of micropropagation, the effects of cytokinin concentrations, and the differences between adventitious and axillary shoots. Greenhouse-grown stock plants were brought into the laboratory, and 4- to 5-cm-long tips of runners were surface-dis-infested for 15 min in 0.5% NaClO with 1 ml of Tween 20/liter, followed by two 5-min rinses in sterile water. Working in the open laboratory near the bases of pairs of lit Bunsen burners, students placed either single-node or shoot tip explants (2 cm long, five replications) onto MS medium with 0, 1, or 10 μM BA. Cultures were in-cubated in parafilm-sealed culture tubes on open laboratory benches. Axillary shoots grew regardless of concentration of BA, and explants on medium with 10 μM BA produced the most callus and adventitious shoots. Microshoots were rooted and ac-climatized under mist in the greenhouse. This exercise can be performed in an open laboratory without the use of laminar flow hoods, specialized sterilizing equipment, or supplemental lighting.

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John E. Preece, Carl A. Huetteman, W. Clark Ashby and Paul L. Roth

Clonal micropropagation studies with silver maple (Acer saccharinum L.) included experiments with various shoot. explant types, cytokinins, and stock plant maturation levels. These trials led to successful explant establishment, axillary shoot proliferation, rooting of microshoots, and establishment of plantlets in the greenhouse. Overall, the best cytokinin tested was the phenylurea derivative TDZ. Shoot proliferation on juvenile explants was poor with kinetin, 2iP, and BA. Only zeatin at 10 μm was comparable to TDZ. TDZ at 10 nm was optimal for both juvenile and adult nodal explants. Juvenile explants that were held in vitro for 4 months commonly had at least 60 axillary shoots that could be subculture or excised for rooting. Microshoots rooted within 2 weeks. Following rooting, silver maple plantlets could be transplanted into a growing medium and placed directly onto a greenhouse bench. Studies were also conducted on rooting stem cuttings (macropropagation). Single nodes from juvenile plants rooted under intermittent mist, regardless of auxin application; however, shoot-tip cuttings from adult trees rooted best when auxin in ethanol solution was applied. Chemical names used: N- phenyl- N' -1,2,3 -thiadiazol-5-ylurea (thidiazuron, TDZ), N- (2-furanylmethyl)-1H-purin-6-amine (kinetin), isopentenyladenine (2iP), benzyladenine (BA), (E)-2-methyl-4-(1H-purin-6-ylamino)-2-buten-1-ol (zeatin).

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John E. Preece, Carl A. Huetteman, W. Clark Ashby and Paul L. Roth

During 1987, we selected the six fastest-growing seedlings or clones from each of 15 provenances that represented the natural distribution range of silver maple (Acer saccharinum L.). Shoots from all 90 trees were cut into nodal segments, rooted as cuttings, and maintained as clonal stock plants in the greenhouse. Rooting was generally excellent and more than half of the clones rooted ≥90%. At the same time, explants were obtained from these field-grown trees and many were established in vitro as aseptic cultures by first pretreating with benomyl and rifampicin. Single-node explants from the greenhouse-grown clonal stock plants were also established and multiplied in vitro. There was a significant effect of clone within provenance on all in vitro growth characteristics. All clones proliferated axillary shoots, but not all at the same rates. Although statistically significant, low correlation coefficients indicated that micropropagation results were not good predictors of nursery performance of the populations from which the clones were selected, nor of the climatic conditions at the site of origin of the trees. The micropropagation system reported herein, therefore, should be applicable to a wide variety of silver maple genotypes. Chemical name used: methyl [1-[butylamino)carbonyl]-1H-benzimidazol-2-yl]carbamate(benomyl).

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John E. Preece, Carl A. Huetteman, W. Clark Ashby, Paul L. Roth and Richard G. Adams

Sixty clones (four clones from each of 15 provenances) were micropropagated and planted in replicated plots in lowland and upland sites in Carbondale, IL in 1991. Data were collected on tree growth, including basal caliper, height, branching, crown volume, dates of bud break, bud set, and leaf fall. There were significant and strong positive genotypic and phenotypic correlations between tree height and basal caliper throughout the three years of growth. During 1993, bud break was not significantly correlated with any growth parameters. After three years in the field, tree height was significantly negatively correlated with the amount of callus that had formed after one month during the in vitro micropropagation phase. However, all shoots that formed in vitro were of axillary origin.

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John E. Preece, Carl A. Huetteman, W. C. Ashby and Paul L. Roth

During the research phase, a system was developed to clonally micropropagate silver maple. Explant performance was best on DKW medium with 10 nM thidiazuron, and explants commonly developed 1 7 shoots after three months and over 60 shoots that could be rooted after four months in vitro. Plants were rooted (>90%) and acclimatized under intermittent mist and transplanted to an outdoor nursery bed. However, results were different during the production phase when 90 clones were propagated. Shoot proliferation rates were lower, differences in clonal response and worker efficiency were apparent, mass rooting under mist was inconsistent and acclimatization problems arose. The mean rooting was 46% under mist because of uneven coverage. Only 56% of rooted plantlets acclimatized which resulted in an overall efficiency of 26%. Partial solutions included root initiation in vitro, and use of fog for acclimatization.