CRISPR (short for “Clustered Regularly Interspaced Short Palindromic Repeats”) is a technology that research scientists use to selectively modify the DNA of living organisms. CRISPR was adapted for use in the laboratory from the naturally occurring genome-editing systems found in bacteria. In this work, we reviewed the methods used to introduce CRISPR/Cas-mediated genome editing into fruit species, as well as the impacts of the application of this technology to activate and knock out target genes in different fruit tree species, including on tree development, yield, fruit quality, and tolerance to biotic and abiotic stresses. The application of this gene-editing technology could allow the development of new generations of fruit crops with improved traits by targeting different genetic segments or even could facilitate the introduction of traits into elite cultivars without changing other traits. However, currently, the scarcity of efficient regeneration and transformation protocols in some species, the fact that many of those procedures are genotype-dependent, and the convenience of segregating the transgenic parts of the CRISPR system represent the main handicaps limiting the potential of genetic editing techniques for fruit trees. Finally, the latest news on the legislation and regulations about the use of plants modified using CRISPR/Cas systems has been also discussed.

Silver nanoparticles (AgNPs) are novel compounds used as antimicrobial and antiviral agents. In addition, AgNPs have been used to improve the growth of different plants, as well as the in vitro multiplication of plant material. In this work the effect of AgNPs on in vitro growth of ‘Canino’ and ‘Mirlo Rojo’ cultivars, as well as the leaf ion composition, are studied. Different concentrations of AgNPs (0, 25, 50, 75 and 100 mg L⁻¹) were added to two culture systems: semisolid medium with agar (SSM) in jars and liquid medium in temporary immersion system (TIS). Proliferation (number of shoots), shoot length, productivity (number of shoot × average length), leaf surface, fresh and dry weight were measured. Additionally, the silver and other ion accumulation in the leaves were evaluated by inductively coupled plasma optical emission spectroscopy (ICP-OES) analysis. The productivity of ‘Canino’ and ‘Mirlo Rojo’ decreased when increasing the concentration of AgNPs in the semisolid medium. However, the use of AgNPs in the TIS improved the proliferation and productivity of ‘Canino’ and Mirlo Rojo’, increasing biomass production, and the concentration of nutrients in the plants, although these effects are genotype-dependent. TISs are the best system for introducing silver into shoots, the optimum concentration being 50 mg L⁻¹ for ‘Canino’ and 75 mg L⁻¹ for ‘Mirlo Rojo’. Principal component analysis, considering all the analyzed ions along the treatments, separates samples in two clear groups related to the culture system used. The use of bioreactors with a liquid medium has improved the productivity of ‘Canino’ and ‘Mirlo Rojo’ in the proliferation stage, avoiding hyperhydration and other disorders. The amount of metallic silver that penetrates apricot plant tissues depends on the culture system, cultivar and concentration of AgNPs added to the culture medium. Silver ion accumulation measured in the shoots grown in the TIS was higher than in shoots micropropagated in a semisolid medium, where it is barely detectable. Furthermore, AgNPs had a beneficial effect on plants grown in TIS. However, AgNPs had a detrimental effect when added to a semisolid medium.

The improvement of previously described protocols for the regeneration of shoots from ‘Canino’ mature seed hypocotyl slices has been accomplished. The effects of different factors such as the part of the hypocotyl used, vacuum-infiltration, 2,4-Dichlorophenoxyacetic acid pulse, vacuum-infiltration and sonication on regeneration and transient transformation were analyzed. When the three slices obtained from the hypocotyls were evaluated separately on regeneration medium, the highest percentages of regenerating explants were achieved in the part close to the epicotyl and in the central part. On the other hand, sonication of the explants for 30 s followed by vacuum-infiltration during Agrobacterium infection for 20 min allowed for an increase in the transformation events. The application of these modifications to the procedure increased the regeneration efficiencies, and transient transformation events and may reduce the frequency of failed experiments. An efficient regeneration/transformation protocol could facilitate its use as a biotechnological technique for apricot breeding.

The use of silver nanoparticles (AgNPs) is increasing nowadays due to their applications against phytopathogens. Temporary Immersion Systems (TIS) allow the micropropagation of plants in liquid media. This work aims to develop an effective protocol for apricot micropropagation in TIS and to study the necessary conditions to introduce AgNPs in apricot plants, as well as the effect of its application on proliferation-related parameters. AgNPs were introduced in different media at a concentration of 100 mg L⁻¹ to test the incorporation of silver to plant tissues. Silver content analysis was made by Inductively Coupled Plasma-Optical Emission Spectroscopy (ICP-OES). The effect of initial shoot density and the addition of AgNPs on micropropagation were evaluated after four weeks in culture on TIS. Productivity, proliferation, shoot-length and leave surface were measured. The better micropropagation rate was obtained with 40 initial shoots, 2 min of immersion every 6 h and 3 min of aeration every 3 h. To introduce AgNPs in apricot plants it is necessary to culture them in liquid media without chloride in its composition. These results will contribute to the development of an in vitro protocol for virus inhibition by AgNPs application. This depends on the introduction of Ag nanoparticles within the plant tissues, and this is not possible if AgNPs after interaction with Cl^- ions precipitate as silver chloride salts.

The methods used to introduce CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats)/Cas-mediated genome editing into fruit species, as well as the impacts of the application of this technology to activate and knock out of target genes in different fruit trees species including tree development, yield, fruit quality, and tolerance to biotic and abiotic stresses have been firstly described in this review. The application of this gene editing technology could allow the development of new generations of fruit crops with improved traits by targeting different genetic segments or even could facilitate the introduction of traits in elite cultivars without changing other traits. However, at this moment, the scarcity of efficient regeneration and transformation protocols in some species, the fact that many of those procedures are the genotype-dependent or the convenience of segregating the transgenic parts of the CRISPR system represent the main handicaps limiting the potential of genetic editing techniques for fruit trees. Finally, latest news on the legislation and regulations about the use of plants modified through CRISPR/Cas systems has been also discussed.

Metallic nanoparticles have antimicrobial, virucidal, and anticancer activities and have been widely applied in medicine. In plants, silver nanoparticles have been used as preventive treatments in the greenhouse to reduce viral titers and symptoms. This work investigates the effect of Argovit™ AgNP formulation on apricot plants infected with Plum pox virus or with Hop stunt viroid. Meristems were rescued from plants treated with different Argovit™ concentrations and two exposure times. Although viroid-free plants were not obtained, a very high efficiency in eliminating the sharka virus is reported with a maximum cleaning efficiency (75 %) after 8 weeks of exposure at 75 mg L^-1 AgNP. To the best of our knowledge, this is the first time that silver nanoparticles application is used, combined with meristem culture, to produce virus-free plants and opens a new path to the elimination of viruses from plants.