Cryopreservation and Acclimatization of <i>Lycopersicon esculentum</i> (Mill.) Genotypes

Abstract

Romanian tomato (Lycopersicon esculentum Mill.) cultivars have been cryopreserved by encapsulation-dehydration and successfully acclimatized to ex vitro growth conditions. Shoot tips were excised from in vitro grown plants then precultured for 24 h in various sucrose concentrations, dehydrated up to 6 h in laminar air flow prior to direct immersion in liquid nitrogen (−196°C) for 24 h. Different parameters have been studied: the effects of osmoprotection and desiccation duration on the regrowth of cryopreserved shoot tips, the effects of various IBA concentrations on rooting and the ex vitro cclimatization of plants recovered from liquid nitrogen. The highest frequency of regrowth (72% cv. ‘Pontica’) was obtained when encapsulated explants were precultured in 0.5 M sucrose and the moisture content (fresh weight basis) of alginate beads was 23%. The highest rooting rates (58% to 77%) for all cultivars were observed for shoots grown on MS medium supplemented with 1.0 mg/l IBA. The rooted plants could be easily acclimatized ex vitro with up to 100% survival.

Managing plant genetic resources using low and ultra-low temperature storage: a case study of tomato

“…It is easy to initiate an aseptic tissue culture from tomato seeds by surface disinfection with 75% Clorox (active chlorine content 5%) for 15 min and rinsing three times with sterile distilled water, followed by germination on a half-strength MS medium, pH 5.7 (Coste et al 2014). A similar protocol, based on surface disinfection of seeds with 3% sodium hypochlorite for 5 min and 70% (v/v) ethanol solution for 30 s, was described by Al-Abdallat et al (2017).…”

“…Therefore, it is recommended to use tomato shoot tips, which are as small as possible, however, big enough to be able to develop after excision and rewarming (FAO 2014). One should also keep in mind, that the cryopreservation efficiency may be cultivar or even genotype specific, as reported by Coste et al (2014Coste et al ( , 2015.…”

“…For example, no shoot recovery was found after using the droplet-vitrification technique (sole pretreatment of tomato shoot tips with PVS2 prior to storage in LN on aluminum foil strips), whereas applying an additional preculture on MS medium with 0.5 M sucrose elevated the results to even 70%. On the other hand, sucrose concentration higher than 0.5 M has an unfavorable effect (osmotic shock) on the species shoot tips survival and recovery rates after cryopreservation, as observed by Coste et al (2014Coste et al ( , 2015.…”

“…Therefore, in order to protect the cells from the adverse effect of concentrated PVS, the tomato explants are first osmoprotected (loading treatment) for 20 min with a loading solution (LS), which is a mixture of low-concentrated, non-penetrating cryoprotectors; usually 2 M glycerol and 1.0 M sucrose (Coste et al 2015;Al-Abdallat et al 2017). Physical (evaporative) dehydration, on the other hand, is performed either over a silica gel or in the stream of (sterile) air flow (Coste et al 2014). The first approach is more rapid and easier to control, while the second one-less shocking and damaging to the tomato tissues.…”

“…The possibility of producing in vitro whole plants from cryopreserved tissues is required. Recovery of cryopreservation-derived tomato shoot tips starts from a few days to about 3 weeks after rewarming, depending on the protocol, with an expansion of the first leaf (Coste et al 2014;Halmagyi et al 2017).…”