Micropropagation, Cutting Propagation and Synthetic Seed Production of Leptospermum Species for Therapeutic Honey Production

Ian Darby

doi:10.25907/00672

Production of Leptospermum seedlings for the establishment of therapeutic-honey plantations has been limited by difficulties in extracting seed from fruit and germinating seed in the nursery. This project aimed to develop alternative methods for micropropagation, cutting propagation and synthetic seed production of Leptospermum plants. Micropropagation studies determined how proliferation and adventitious rooting of Leptospermum polygalifolium and L. scoparium shoots were influenced by the concentration of benzyladenine (BA) in the proliferation medium and the original explant position on the seedling. Hormone-free node culture was highly effective for both species. Nodal explants often formed roots in the absence of BA and developed elongated axillary shoots. Median shoot numbers of 584 and 659 were formed within 32 weeks from a single L. polygalifolium or L. scoparium seed, respectively. A low BA dose was effective for callogenesis and shoot proliferation of L. polygalifolium, but not L. scoparium. The median number of shoots produced from a single L. polygalifolium seed using 2.22 μM BA was 630. This BA dose induced extremely high shoot numbers in some clones because explants often produced extensive callus and multiple short shoots. Shoots formed adventitious roots without indole-3-butyric acid and plantlets were acclimatised successfully to nursery conditions. The original explant position did not influence shoot proliferation or adventitious rooting. Leptospermum polygalifolium and L. scoparium proved amenable to micropropagation, facilitating rapid establishment of nectar plantations. Stock plants were produced in the nursery from the micropropagated L. polygalifolium and L. scoparium plantlets. We assessed the amenability of these two high-value species to propagation as rooted cuttings with or without application of the rooting hormone, indole-3-butyric acid (IBA). IBA did not significantly affect average rooting percentages, although it occasionally increased the number of adventitious roots per rooted cutting. Average rooting percentages for untreated cuttings were 92%, 88% and 93% for three L. polygalifolium clones and 65% and 76% for two L. scoparium clones. The cumulative capacity for plant production from three successive harvests of cuttings was 95, 126 and 118 rooted cuttings per stock plant for the L. polygalifolium clones and 37 and 53 rooted cuttings per stock plant for the L. scoparium clones. Leptospermum polygalifolium and L. scoparium were both highly amenable to vegetative propagation, again facilitating the establishment of nectar plantations for producing therapeutic honey. Shoots of L. polygalifolium and L. scoparium that were produced by micropropagation were also used to assess the amenability of these species to propagation and germplasm distribution using synthetic seeds. This study aimed to determine how nutrient strength of the encapsulation solution and presence of BA in the emergence medium affected shoot and root emergence from synthetic seeds. Nodes from in vitro shoots were encapsulated in 3% sodium alginate with either half- or full-strength Murashige and Skoog (MS) medium and the synthetic seeds were placed on emergence medium containing either 0 or 2.2 μM BA. Full-strength MS was effective for shoot emergence of both species. Half-strength MS was insufficient for shoot emergence from L. polygalifolium but allowed shoot emergence from many L. scoparium synthetic seeds. BA increased the percentage of synthetic seeds with shoot emergence in one L. polygalifolium clone, but it reduced the percentage of synthetic seeds with shoot emergence in one L. scoparium clone. BA sometimes stimulated shoot emergence through callus in both species. Furthermore, BA increased the number of shoots per emergent synthetic seed of L. scoparium. Surprisingly, the simple use of full-strength MS in the encapsulation solution, without BA in the emergence medium, stimulated root emergence from 25–58% of L. polygalifolium and 100% of L. scoparium synthetic seeds. These results demonstrate that L. polygalifolium and L. scoparium are highly amenable to propagation and distribution as synthetic seeds, which could assist greatly in mass-production of plants for therapeutic-honey plantations. This research has developed novel techniques for the production of shoots, plantlets, cuttings and synthetic seeds of L. polygalifolium and L. scoparium for nursery plant production and plantation establishment. These techniques offer alternatives to the traditional, difficult, seedling-propagation methods for Leptospermum species. They allow the mass-production of two high-value species for nectar plantations, which will underpin production of therapeutic honey with antimicrobial and wound-healing properties.

Micropropagation, Cutting Propagation and Synthetic Seed Production of Leptospermum Species for Therapeutic Honey Production

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