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Rapid in vitro shoot multiplication of the recalcitrant species Juglans nigra L.

  • Micropropagation
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Abstract

Black walnut (Juglans nigra L.) has long been prized for its timber, leading to commercial cultivation and significant breeding efforts for improving marketable traits. Vegetative and in vitro black walnut propagation techniques, however, are variable and highly genotype dependent. Optimizing plant growth regulator type and concentration are integral for developing a successful micropropagation protocol. The addition of meta-topolin (MT) combined with the novel use of a liquid medium has led to the development of a rapid shoot multiplication system. The objective of this research was to develop a reproducible and dependable micropropagation protocol for elite black walnut genotypes. In vitro shoot cultures were established from nodal explants cultured on semi-solid Driver and Kuniyuki walnut (DKW) medium supplemented with 8.9 μM benzyladenine, 0.005 μM indole-3-butyric acid (IBA), 200 mg L−1 casein hydrolysate, 50 mg L−1 adenine hemisulfate, 2 mL L−1 Plant Preservative Mixture™, and 4.1 μM MT. Long-term survival and proliferation of microshoots was achieved when nodal segments of in vitro grown shoots were cultured in liquid initiation medium in 3-L polycarbonate Fernbach-style flasks on a rotary shaker (100 rpm) under a 16-h photoperiod at 25°C. Elongated microshoots (5–7 cm in length) were rooted in a slurry-like medium composed of half-strength DKW medium with 0.11% (w/v) Phytagel™ and coarse vermiculite (2:1, v/v) supplemented with 50 μM IBA for 5 wk. Rooted shoots were acclimatized to ambient culture room conditions, but plantlets did not survive once transferred to the greenhouse.

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Funding

Financial support for this work came from a Fred M. van Eck scholarship for Purdue University to Micah Stevens. The mention of a trademark, proprietary product, or vendor does not constitute a guarantee or warranty of the product by the U.S. Dept. of Agriculture and does not imply its approval to the exclusion of other products or vendors that also may be suitable.

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Authors and Affiliations

  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

    Micah E. Stevens

  2. USDA Forest Service, Northern Research Station, HTIRC, 715 West State St, West Lafayette, IN, 47907, USA

    Paula M. Pijut

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  1. Micah E. Stevens
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  2. Paula M. Pijut
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Correspondence to Paula M. Pijut.

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Editor: Wenhao Dai

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Stevens, M.E., Pijut, P.M. Rapid in vitro shoot multiplication of the recalcitrant species Juglans nigra L.. In Vitro Cell.Dev.Biol.-Plant 54, 309–317 (2018). https://doi.org/10.1007/s11627-018-9892-3

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