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Direct and indirect in vitro regeneration of Miscanthus × giganteus cultivar Freedom: effects of explant type and medium on regeneration efficiency

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Abstract

Miscanthus × giganteus (giant miscanthus; Mxg) is a seed-sterile, perennial bioenergy crop with the potential to produce liquid fuel from lignocellulosic biomass. A new cultivar, Freedom, is being commercially grown in the USA on increasing acreage. To determine this genotype’s regeneration responses in tissue culture, three explant sources were screened on media proven successful for other genotypes. Four callus induction media contained 13.6–22.6 μM 2,4-dichlorophenoxyacetic acid (2,4-D) alone or with 0.44–4.4 μM 6-benzyladenine (BA). Callus induction percentages for all explants ranged from 93 to 97%. Media yielding the greatest percentages of explants producing regenerable calli for shoot apices (from in vitro and greenhouse plant sources) were media containing either 13.6 μM 2,4-D plus 0.44 μM BA or 22.6 μM 2,4-D plus 0.44 μM BA. After culture on a regeneration medium containing 22 μM BA plus 1.3 μM naphthaleneacetic acid (NAA), 3.59–3.74 regenerants were obtained per explant. Immature inflorescence explants (from field-maintained plants) gave up to 77% regenerable calli and 6.99 regenerants per explant. Direct regenerants (shoots) arose from immature inflorescence explants on a medium containing 9.0 μM 2,4-D. Intact plants could be generated within 16–18 wk after culture initiation. Extensive visual assessments, and molecular assessments via inter-simple sequence repeat (ISSR) PCR analysis using 21 different primers, did not reveal distinguishable somaclonal variation among regenerants or when compared to rhizome-propagated transplants under field conditions. We believe that this is the first extensive in vitro and ex vitro analysis on a commercially grown Mxg genotype.

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Acknowledgments

This research was funded by Repreve Renewables Inc., Soperton, GA, and underwritten by the US Department of Energy. The authors thank Dr. Karen Koefoed Petersen, Danish Institute of Plant and Soil Sciences, Denmark, for providing valuable expertise and advice. We also thank Dr. Richard Harkess, Department of Plant and Soil Sciences, and Dr. Maria Tomaso-Peterson, Department of Biochemistry, Molecular Biology, Entomology, and Plant Pathology, both from Mississippi State University, for providing editorial comments on previous drafts.

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Correspondence to Dinum Perera.

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Editor: Baochum Li

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Perera, D., Barnes, D.J., Baldwin, B.S. et al. Direct and indirect in vitro regeneration of Miscanthus × giganteus cultivar Freedom: effects of explant type and medium on regeneration efficiency. In Vitro Cell.Dev.Biol.-Plant 51, 294–302 (2015). https://doi.org/10.1007/s11627-015-9682-0

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