Microspore embryogenesis is a method of achieving complete homozygosity from plants. It is particularly useful for woody species, like Citrus, characterized by long juvenility, a high degree of heterozygosity and often self-incompatibility. Anther culture is currently the method of choice for microspore embryogenesis in many crops. However, isolated microspore culture is a better way to investigate the processes at the cellular, physiological, biochemical, and molecular levels as it avoids the influence of somatic anther tissue. To exploit the potential of this technique, it is important to separate the key factors affecting the process and, among them, culture medium composition and particularly the plant growth regulators and their concentration, as they can greatly enhance regeneration efficiency. To our knowledge, the ability of meta-Topolin, a naturally occurring aromatic cytokinin, to induce gametic embryogenesis in isolated microspores of Citrus has never been investigated. In this study, the effect of two concentrations of meta-Topolin instead of benzyladenine or zeatin in the culture medium was investigated in isolated microspore culture of two genotypes of Citrus. After 11 months of isolated microspore culture, for both genotypes and for all the four tested media, the microspore reprogramming and their sporophytic development was observed by the presence of multinucleated calli and microspore-derived embryos at different stages. Microsatellite analysis of parental and embryo samples was performed to determine the embryo alleles constitution of early embryos produced in all tested media, confirming their origin from microspores. To our knowledge, this is the first successful report of Citrus microspore embryogenesis with isolated microspore culture in Citrus, and in particular in Citrus clementina Hort. ex Tan, cvs. ‘Monreal Rosso’ and ‘Nules.’

Three caper (Capparis spinosa L.) biotypes grown on the Sicilian island of Salina (38°33′49″ N) were micropropagated to evaluate two different in vitro culture systems: one using the traditional solid medium, and the other based on liquid culture in a PlantForm bioreactor. PlantForm is a temporary immersion system (TIS), a new propagation method in which the shoots undergo temporary immersion in a liquid medium in order to avoid the accumulation of gas through forced ventilation. This study proposes a protocol to improve the efficiency of in vitro propagation of caper plants, while also reducing production costs, because of the elimination of the gelling agent, and manual labor, requiring limited subcultures and posing minimal contamination risks. Our results show that the caper shoots propagated in bioreactors demonstrated good adaptability and better growth rates than those grown in the conventional system. Statistically significant differences were observed between plants grown in the PlantForm liquid culture and those grown in solid medium regarding the number and length of shoots, which were further promoted by the addition of plant growth regulators (PGRs). The relative growth and real proliferation rate of the caper explants were higher when using meta-Topolin than when using 6-benzylaminopurine as a PGR. Overall, the TIS improved in vitro caper culture by promoting the proliferation, length, and vigor of the shoots.

The in vitro rooting of three caper (Capparis spinosa L.) selected biotypes, grown in a commercial orchard on the Sicilian island of Salina (38°33′49” N), was performed using—as base material for rooting experiments—shoot explants proceeding from two different in vitro culture systems: solid medium and liquid culture in a PlantForm bioreactor (TIS). The regenerated shoots of each accession were submitted to different auxin treatments (NAA, IBA, IAA - 1 or 2 mg L−1; NAA+IBA 0.75 and 0.25 mg L−1, respectively), supplemented with sucrose or fructose (mg L−1). The highest rooting rate in terms of root percentage (67%) was reached with the explants of the selected accession ‘Sal 39’ proceeding from liquid culture in PlantForm and induced in the MS medium with sucrose, as a carbon source, supplemented with NAA 0.75 mg L−1 + IBA 0.25 mg L−1, after six days in a climatic growth chamber at 25 ± 1 °C in the dark and then placed under a cool white fluorescent lamp, with a PPFD of 35 μmol m−1 s−1 and a photoperiod of 16 h. On the other hand, poor rooting rate was generally achieved under all the tested experimental conditions with the other biotypes, ‘Sal 37’ and ‘Sal 35’, demonstrating the strong role exerted by the previously adopted proliferation method and by the genotype for successful caper in vitro rooting.

No abstract

No abstract