Progress 09/01/00 to 08/31/04
Outputs
The North American Pawpaw shows great potential as a new fruit crop. Kentucky State University in Frankfort, Ky. is the site for the USDA National Clonal Germplasm Repository for Asimina species. Both the fruit and the trees themselves are of high value to growers and nursery producers. Pawpaw cultivars are currently propagated by grafting or budding onto seedling rootstock; no method currently exists to clonally propagate pawpaw on its own roots. Three methods of layering were attempted in this study to clonally propagate pawpaw: trench layering, pot layering, and mound layering. Both trench layering and pot layering experiments showed the importance of both juvenility and auxin application in adventitious rooting of pawpaw. Although rooting of more mature pawpaw shoots in these experiments did not exceed 30%, these propagation methods were more successful then previous attempts at rooting more mature pawpaw stems. Mound layering was less successful, but an
easier-to-root genotype of pawpaw in the KSU-USDA repository orchards was discovered that shows promise for future propagation studies. Efforts to mass propagate pawpaw in vitro met with limited success. The ability for single stem explants to produce shoots from 3-year-old cultures of A10-11 was investigated using medium containing 9.8 uM IBA plus 5.4 uM NAA in combination with BA (0 to 20 uM). Initial explants elongated but did not form additional shoots after 8 weeks in culture. These were subcultured to the same medium and after 9 weeks cultures treated with 15 or 20 uM BA. The 15 uM BA treatment had the most vigorous shoot growth and shoots per culture. These data indicate that cultures of pawpaw can retain morphogenetic potential for a considerable time in culture. Efforts to root shoots in vitro were unsuccessful. Pawpaw microshoots of selection A10-11 that were established in culture 6 years previously now continue to proliferate without the addition of hormones to the culture
medium. This phenomenon, called habituation, has been observed in callus culture of other plants, but never before with whole organs (microshoots) in tissue culture.
Impacts
Methods for the clonal propagation of pawpaw were developed that can be utilized by nurseries and growers. Up to 30% of mature pawpaw stems rooted in trench and pot layering methods; these rooting percentages are far more successful than with previous attempts at rooting mature pawpaw stems. These layering propagation methods can be utilized to clonally propagate pawpaw on its own roots. Methods to enhance pawpaw shoot production in vitro have shown promise; however, there is currently no method to root shoots produced in vitro.
Publications
- Pomper, K.W. and D.R. Layne. 2005. The North American Pawpaw: Botany and Horticulture. Hort. Rev. Vol. 31:351-384.
- Crabtree, S.B. 2004. Sexual and Asexual Reproductive Characteristics of the North American Pawpaw [Asimina triloba (L.) Dunal]. Master of Science Thesis. University of Kentucky, Department of Horticulture.
- Pomper, K.W. and D.R. Layne. 2004. North American Pawpaw. Chronica 44:11-15.
- Sheri B. Crabtree, Kirk W. Pomper, and Robert L. Geneve. 2004. Trench layering as a method of clonally propagating pawpaw (Asimina triloba). HortScience 39:890
- Geneve, R. L., K. W. Pomper, S. T. Kester, J. N. Egilla, C. L. H. Finneseth, S. B. Crabtree, and D. R. Layne. 2003. Propagation of pawpaw - a review. HortTechnology 13: 428-433.
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Progress 01/01/03 to 12/31/03
Outputs
The pawpaw is the largest fruit native to the U.S. and has potential as a new fruit crop. Few methods are available to clonally propagate pawpaw, with grafting or budding onto a seedling rootstock being the only currently feasible method. Developing new options for clonally propagating pawpaw could facilitate the development of a commercial industry. Trench layering was examined as a method to clonally propagate pawpaw. In a greenhouse factorial experiment the influence of plant juvenility or age and auxin concentration on rooting was examined. Seedlings that had emerged 3, 6, and 12 weeks prior to the experiment were defoliated, tipped, and transplanted into greenhouse beds. Shoots were etiolated, then girdled and treated with either 0, 5000, or 10,000 ppm IBA (auxin). The main effects of age and auxin concentration significantly affected the percentage of shoots producing roots. More juvenile or younger plants had an increased ability to produce roots, with 15% of
the shoots of the 3-week old seedlings producing at least one root, compared to only about 5% of the 12 week old seedlings rooting. Auxin application to shoots also promoted rooting, with 16% of IBA-treated shoots producing at least one root, compared to the untreated control, with only 2% of shoots producing a root. There was no significant difference in rooting percentage between the two concentrations of IBA. The treatment combination that produced the greatest number of shoots with at least one root was 10,000 ppm IBA applied to shoots of 3 week old seedlings, with 31% of shoots rooting. Experiments to develop a micropropagation protocol for pawpaw have been unsuccessful; however, experiments to promote shoot proliferation and elongation are continuing.
Impacts
The inability to easily clonally propagate pawpaw has been a major factor limiting development of a commercial pawpaw industry. Methods to enhance pawpaw shoot production in tissue culture have been pursued. Attempts to utilize mound layering to clonally propagate pawpaw have had limited success; however, trench layering has great potential as a propagation method for pawpaw.
Publications
- Geneve, R. L., K. W. Pomper, S. T. Kester, J. N. Egilla, C. L. H. Finneseth, S. B. Crabtree, and D. R. Layne. 2003. Propagation of pawpaw - a review. HortTechnology 13: 428-433.
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Progress 01/01/02 to 12/31/02
Outputs
Developing a standard micropropagation protocol for pawpaw, requires the ability to consistently produce shoot microcuttings from various sources of plant material. A shoot growth response curve was developed for two cytokinins, and the effect of auxin versus cytokinin on shoot growth of micropropagules in vitro was characterized. To determine the effects of two cytokinins on biomass production, micropropagules with multiple shoots were transferred onto an agar-solidified Murashige and Skoog (MS) basal salt medium containing the auxins: indole-3-butyric acid (IBA) (9.8 uM) and naphthalene acetic acid (NAA) (5.37 uM) and the cytokinins: 6-benzyladenine (BA) or kinetin (0, 5, 10, 15 and 20 uM). Data obtained from this study indicated that optimum BA or kinetin for maximum shoot biomass production of pawpaw was about 15 uM. BA promoted greater biomass production compared to kinetin. Shoot elongation was achieved only at 5 uM BA, but multiple bud production required
higher concentrations of BA and kinetin. Additionally, relative growth rate (RGR) was determined as daily increase in fresh mass at two temperature regimes during the last 38 days in culture prior to termination. Increasing the culture temperature by 3 C (28.6 vs. 25.5 C) significantly increased biomass and RGR during the incubation period in cultures containing BA, but not kinetin. To examine whether mound layering could be used to clonally propagate pawpaw, a factorial experiment was conducted with two levels of girdling (girdled or not) and three levels of IBA at 0, 3000, and 6000 ppm in lanolin paste. Eight trees were selected in May 2002, cut back to 10 cm above the ground, and covered with sawdust to cause etiolation of the new shoots. Treatments were applied on July 15, 2002 to actively growing shoots. Data on the number of surviving shoots, number of shoots producing roots, number of roots per shoot, root length, and shoot diameter were recorded in November, 2002. Only one
shoot from one tree in all treatments had a root develop.
Impacts
Difficulties in the propagation of pawpaw have been a major factor limiting development of a commercial pawpaw industry. Methods to enhance pawpaw shoot production in tissue culture are being pursued. Initial attempts to utilize mound layering to clonally propagate pawpaw were unsuccessful; however, we are examining a modified layering approach to promote root formation.
Publications
- No publications reported this period
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Progress 01/01/01 to 12/31/01
Outputs
Two experiments were conducted to develop a feasible method for in vitro propagation of the North American pawpaw (Asimina spp.). Experiment 1 involved the aseptic establishment of two pawpaw species (Asimina longifolia Kral. and Asimina tetramera Small) in culture. In this Stage 1 of the in vitro culture process, explants from nodal segments of shoots from the two pawpaw species were established aseptically in Murashige and Skoog (MS) basal medium containing 2 uM IBA, 1 uM NAA, 1 uM Kinetin, and 10 uM BA, and 0.7% Agar. All surviving explants were transferred into a fresh medium containing the same concentrations of auxin and cytokinin four weeks after stage 1 was initiated. Initial and final shoot lengths of the explants were recorded during subculture and at termination of experiment, respectively. In experiment 2, the effect of benzyl adenine (BA) on the growth rate of pawpaw [Asimina triloba (L.) Dunal] microshoots previously developed from in vitro was evaluated
in McCown's Woody Plant Medium (WPM) containing 1.0 mg/L (5.37 uM NAA), 2.0 mg/L (9.8 uM IBA), and four concentrations of BA (0, 5, 10, 15 and 20 uM). Culture tubes were incubated in a Growth Chamber (G/C) at 25 oC, under uninterrupted lighting with cool white fluorescent lamp at approximately 16.5 umol m-2 s-1. Microshoots were observed weekly for mortality and total number of culture tubes with contamination. After 8 weeks of incubation in the G/C, the number of adventitious shoots produced, and increase in shoot length of each microshoot were quantified. From the data, explants in the MS medium without auxin or cytokinin [(-) PGR] had higher percent survival (40%) than explants cultured in the same MS medium with both auxin and cytokinin [(+) PGR] (5%). All the A. tetramera explants established in aseptic culture died three days after the initiation of experiment 1. In experiment 2, based on the percent survival and the mean increase in shoot length in vitro, the 15 uM BA
concentration promoted the most microshoot development during the eight-week culture period at 16.5 umol m-2s-1 among the BA concentrations tested. To examine if mound layering could be used to clonally propagate pawpaw (A. triloba), a factorial experiment was conducted with two levels of girdling (girdled or not girdled) and three levels of IBA at 0, 3000, and 6000 ppm in lanolin paste. Ten trees were selected in early spring and cut back to about 10 cm above the ground, and treatments applied in early summer to the actively growing semi-hardwood shoots (each tree served as a block containing all treatments). Root production on shoots will be evaluated next spring.
Impacts
Difficulties in the propagation of pawpaw have been a major factor limiting development of a commercial pawpaw industry. Methods to establish pawpaw shoots and promote shoot growth in tissue culture were partially successful. Initial attempts to utilize mound layering to clonally propagate pawpaw were not successful; however, we are attempting a modified approach using this technique.
Publications
- No publications reported this period
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