Hypocotyl derived in vitro regeneration of pumpkin ash (<Emphasis Type="Italic">Fraxinus profunda</Emphasis>) |
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Authors: | Micah E Stevens Paula M Pijut |
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Institution: | (1) Department of Forestry and Natural Resources, Hardwood Tree Improvement and Regeneration Center (HTIRC), Purdue University, 715 West State St., West Lafayette, IN 47907, USA;(2) USDA Forest Service, Northern Research Station, HTIRC, 715 West State St., West Lafayette, IN 47907, USA; |
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Abstract: | Pumpkin ash (Fraxinus profunda (Bush) Bush) is at risk for extirpation by an exotic insect, the emerald ash borer (EAB). Pumpkin ash is limited to wetland
areas of the Eastern United States, and has been listed as an endangered species because of EAB activity. Pumpkin ash provides
many benefits to the ecosystem, and its wood is used in the manufacturing industry. In vitro regeneration provides an integral
tool for the mass propagation and genetic transformation of pumpkin ash to combat EAB. Therefore, a plant regeneration protocol
was developed for pumpkin ash. Aseptically extracted hypocotyls formed adventitious shoots following 4 weeks on Murashige
and Skoog (MS) medium supplemented with 0–22.2 μM 6-benzyladenine (BA) and 0–6.8 μM thidiazuron (TDZ) then transferred for
an additional 4 weeks on MS medium with Gamborg B5 vitamins plus 0.2 g L−1 glycine (B5G) containing 6.7 μM BA, 1 μM indole-3-butryic acid (IBA), and 0.29 μM gibberellic acid (GA3). As adventitious shoots developed, these were transferred to a MSB5G medium with 13.3 μM BA, 1 μM IBA, and 0.29 μM GA3 for shoot elongation. Elongated shoots were successfully micropropagated using MSB5 medium with 10 μM BA and 10 μM TDZ. Adventitious
root formation was as high as 94% using woody plant medium supplemented with 4.9 μM IBA with shoots cultured for 10 days in
the dark followed by culture under a 16-h photoperiod. Acclimatization to the greenhouse was successful and normal plant growth
was observed. This protocol will provide a means for genetic transformation for EAB resistance and mass propagation for conservation. |
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