No abstract

Small non-coding RNAs (sncRNAs) are molecules with important regulatory functions during development and environmental responses across all groups of terrestrial plants. In seed plants, the development of a mature embryo from the zygote follows a synchronized cell division sequence, and growth and differentiation events regulated by highly regulated gene expression. However, given the distinct features of the initial stages of embryogenesis in gymnosperms and angiosperms, it is relevant to investigate to what extent such differences emerge from differential regulation mediated by sncRNAs. Within these, the microRNAs (miRNAs) are the best characterized class, and while many miRNAs are conserved and significantly represented across angiosperms and other seed plants during embryogenesis, some miRNA families are specific to some plant lineages. Being a model to study zygotic embryogenesis and a relevant biotechnological tool, we systematized the current knowledge on the presence and characterization of miRNAs in somatic embryogenesis (SE) of seed plants, pinpointing the miRNAs that have been reported to be associated with SE in angiosperm and gymnosperm species. We start by conducting an overview of sncRNA expression profiles in the embryonic tissues of seed plants. We then highlight the miRNAs described as being involved in the different stages of the SE process, from its induction to the full maturation of the somatic embryos, adding references to zygotic embryogenesis when relevant, as a contribution towards a better understanding of miRNA-mediated regulation of SE.

No abstract

Plant regeneration is a well-known capacity of plants occurring either in vivo or in vitro. This potential is the basis for plant micropropagation and genetic transformation as well as a useful system to analyse different aspects of plant development. Recent studies have proven that RNA species with no protein-coding capacity are key regulators of cellular function and essential for cell reprogramming. In this review, the current knowledge on the role of several ncRNAs in plant regeneration processes is summarized, with a focus on cell fate reprogramming. Moreover, the involvement/impact of microRNAs (miRNAs), long non-coding RNAs (lncRNAs) and small-interfering RNAs (siRNAs) in the regulatory networks of cell dedifferentiation, proliferation and differentiation is also analysed. A deeper understanding of plant ncRNAs in somatic cell reprogramming will allow a better modulation of in vitro regeneration processes such as organogenesis and somatic embryogenesis.

Somatic embryogenesis in Solanum betaceum (tamarillo) has proven to be an effective model system for studying morphogenesis, since optimized plant regeneration protocols are available, and embryogenic competent cell lines can be induced from different explants. Nevertheless, an efficient genetic transformation system for embryogenic callus (EC) has not yet been implemented for this species. Here, an optimized faster protocol of genetic transformation using Agrobacterium tumefaciens is described for EC. The sensitivity of EC to three antibiotics was determined, and kanamycin proved to be the best selective agent for tamarillo callus. Two Agrobacterium strains, EHA105 and LBA4404, both harboring the p35SGUSINT plasmid, carrying the reporter gene for β-glucuronidase (gus) and the marker gene neomycin phosphotransferase (nptII), were used to test the efficiency of the process. To increase the success of the genetic transformation, a cold-shock treatment, coconut water, polyvinylpyrrolidone and an appropriate selection schedule based on antibiotic resistance were employed. The genetic transformation was evaluated by GUS assay and PCR-based techniques, and a 100% efficiency rate was confirmed in the kanamycin-resistant EC clumps. Genetic transformation with the EHA105 strain resulted in higher values for gus insertion in the genome. The protocol presented provides a useful tool for functional gene analysis and biotechnology approaches.