Optimizing micropropagation conditions for a recalcitrant ninebark (Physocarpus opulifolius L. maxim.) cultivar

Abstract

Ninebark is a very popular ornamental shrub. Micropropagation is an efficient method for mass production of uniform plant material. This study was designed to develop and optimize conditions at all phases of ninebark micropropagation. For the multiplication stage, the Murashige and Skoog (MS) medium at full concentration and pH 5.8 was chosen as the basal medium. Sorbitol proved a more effective carbohydrate source than fructose, with no adverse effects on shoot vitrification or the medium itself. The best shoot production, both in number and length, was on the medium enriched with 2 and 3 mg·L−1 zeatin. High numbers of shoots were also obtained in treatments with 1 mg·L−1 6-benzyladenine (BA) or 2 mg·L−1 meta-Topolin (mT) in the basal medium. BA was the most cost-effective cytokinin. There was a positive effect of the gibberellic acid on proliferation: the highest shoot number per explant was produced in the presence of 1 mg·L−1 GA3. No effect of the culture age (up to 20 subcultures) on the percentage of regenerating explants was evident, and the highest numbers of shoots were obtained between passages 10 and 17. For rooting, the MS medium at half strength was used. The best rooting was at 1 mg·L−1 IBA. Spraying the in vitro rooted cuttings with abscisic acid (ABA) favored plant acclimation to the ex vitro conditions. Exvitro rooting, including the treatments with IBA and ABA, shortened the production time by approximately one third.

High-Frequency Plant Regeneration, Genetic Uniformity, and Flow Cytometric Analysis of Regenerants in Rutachalepensis L.

“…Sucrose was shown to be the most effective sugar for promoting shoot regeneration from nodal explants in this study, followed by fructose, maltose, and glucose. Similar results were obtained in micropropagation of Harpagophytum procumbens [56], Aquilaria malaccensis [95], Plumbago zeylanica [91], Pterocarpus marsupium [96], and Lupinus albus [97]. Sucrose, the most prevalent carbohydrate in plant phloem sap [50,51], is often used in tissue culture due to its inexpensive cost, easy availability, and ability to quickly absorb through the plasma membrane [74], which aids in sugar hydrolysis as well as maintain the pH of the medium [48].…”

“…However, shoot differentiation and growth were found less effective in B5, WPM, NN, and White media with a pH 5.8. MS medium with a pH 5.8 has also been found significantly more effective than other media in various plants species, e.g., Albizia lebbeck [26], Vitex negundo [25], Centella asiatica [27], Plumbago zeylanica [91], and Physocarpus opulifolius [92].…”

Acclimation to Ex Vitro Conditions in Ninebark

“…All experiments were done on the red ninebark (Physocarpus opulifolius L.) cultivar Diable d'Or Mindia. The initial material came from 8-10-week-old in vitro culture growing on 1 /2MS supplemented with 1 mg•L −1 IBA [13]. Plants were at least 3.5 cm high, on average had 9 internodes and 4-6 roots not shorter than 1 cm.…”

Morpho-anatomical and biochemical changes associated with rooting of micropropagated ninebark cuttings

“…The experimental material were the apical segments of shoots collected from established 8-10 week old in vitro cultures of ninebark cv. DIABLE D'OR 'Mindia' growing on the MS medium (Murashige and Skoog 1962) with 1 mg•L −1 BA and 0.1 mg•L −1 NAA (Jagiello- Kubiec et al 2021). Sorbitol was used as the carbon source as it proved more suitable than fructose (Jagiello-Kubiec et al 2021).…”