Gametic and somatic embryogenesis through in vitro anther culture of different Citrus genotypes

Abstract

In vitro tissue culture represents a useful technique for advancing Citrus breeding and propagation. Among in vitro regeneration systems, anther culture is commonly used to produce haploids and doubled haploids for a fast-track producing homozygous lines, in comparison with the traditional self-pollination approach, which involves several generations of selfing. In addition, anthers culture can produce somatic embryos that can also be used for clonal propagation. In this study, two thermal shocks were applied to the anthers of six Citrus genotypes (two clementine and four sweet oranges), just after they were put in culture. The response obtained was different depending on the genotype: both clementines, namely Hernandina and Corsica, produced homozygous and triploid regenerants (microspore-derived embryos), whereas all of the analyzed regenerants from sweet oranges, three cultivars of Tarocco and Moro, produced heterozygous and diploid regenerants similar to the parental genotypes (somatic embryos).

Ploidy manipulation for genetic improvement in some Mediterranean fruit crops

“…For this reason, they were considered diploids. Actually, through this technique, it is possible to obtain heterozygous diploid embryoids from the somatic tissue of the anther (Cardoso et al 2016). In fact, anther culture is often used to achieve somatic embryos, since they can be employed for clonal propagation, synthetic seed production, protoplast fusion or genetic transformation (Germanà 2005;Germanà et al 2011a, b).…”

“…Statistical analysis of variance (ANOVA, Tukey's test) showed, as expected, a marked and significant (p ≤ 0.05) genotype effect for all parameters. Concerning the culture media, P and N6 media were previously tested in many woody plant species, such as apricot, citrus, loquat and hazelnut, to induce gametic embryogenesis (Germanà et al 1996;Germanà and Chiancone 2003;Karasawa et al 2016;Cardoso et al 2016). In our case, a significant (p ≤ 0.05) culture medium effect was observed (especially in the percentage of production of calli and embryos), with the exception of undeveloped anthers.…”

“…In particular, for both registered parameters (AA: androgenic anthers and MC: morphogenic calli), the highest percentages (19.3 and 12.7 respectively) were detected in 'Sanfilippara', while the lowest percentages (1.94 and 0.97) were observed in 'Redonet'. In anthers culture, the effect of type and concentration of plant growth regulators has been widely studied (Oleszczuk et al 2004;Perera et al 2009;Smýkalová et al 2009;Germanà 2011a;Cardoso et al 2016). In woody species, different auxin types and concentrations, such as NAA, TDZ or 2.4-D, have been already employed to induce callus production in mandarin (Germanà and Chiancone 2003), papaya (Rimberia et al 2007) and also in loquat (Li et al 2008;Blasco et al 2015b).…”

“…In fact, induction protocols vary not only among species but also among genotypes of the same species. Recently, species previously considered recalcitrant, such as woody plants (Germanà 2006(Germanà , 2009Chiancone and Germanà 2016;Cardoso et al 2016), have been objects of successful studies, causing a resurgence of interest in Hs and DHs of higher plants (Forster et al 2007a;Ferrie 2009;Wedzony et al 2009;Dunwell 2010;Ferrie and Caswell 2011;Islam and Tuteja 2012). In addition, there is a strong commercial interest in methods for the production and exploitation of H and DH plants, verified by the extensive application of intellectual property protection and patenting system (Dunwell 2009(Dunwell , 2010).…”

“…'Hamlin' (C. clementina × C. sinensis) (Cardoso et al 2014), homozygous microspore-derived embryos from cvs. 'Monreal Rosso' and 'Nules' (Chiancone et al 2015) and homozygous and triploid regenerants of 'Hernandina' and 'Corsica' clementines (Cardoso et al 2016), were also obtained.…”