Genetic stability of plants during in vitro propagation and conservation is one of the important aspects of plant biotechnology. In the present study, micropropagated P. lentiscus L. shoot cultures, which are cultivated for the mastic resin, have been cold stored up to 12 months at 4°C in the dark for different durations (2, 4, 6, 8, 10 and 12 months) and genetic alterations in cold storage conditions were evaluated. Growth parameters such as proliferation rate, shoot numbers per explant, shoot lengths and shoot forming capacity were also calculated. Since the highest proliferation rate (100 %) was obtained in 6 month-stored shoot cultures without any severe influence of cold stress on proliferation ability, amplified fragment length polymorphism (AFLP) and inter-retrotransposon amplified polymorphism (IRAP) marker systems were used to determine genetic stability in the plantlets after this storage period. Totally, 702 scorable bands were produced by 10 AFLP primer pairs. Genetic similarity value of the non-stored (control) plant and coldstored clones ranged from 0.66 to 0.84 with a mean of 0.74. In the case of IRAP, 159 bands were produced by 8 IRAP primers. Genetic similarity value of the non-stored plant and cold-stored clones varied from 0.65 to 0.83 and the average genetic similarity value was determined as 0.72. The genetic similarity indices revealed that genetic variability was similar in both techniques. Our results showed that tissue culture and especially cold storage of P. lentiscus L. may result transposons activation, thus could cause genetic instability.
Different nutrient media (MS [Murashige and Skoog 1962]; QL [Quoirin and Lepoivre 1977] and WPM [Lloyd and McCown 1980]); plant growth regulators BA (benzil adenin), GA3 (gibberellic acid), IBA (indole-3-butyric-acid), NAA (naftalen acedic acid); and sucrose concentrations were studied to determine the in vitro culture effects on healthier and faster seedling development from mature lentisk (Pistacia lentiscus L.) seeds. After 28 days of culture, the percentage of germinated seeds was the highest (70%) in the full-strength MS medium. The cytokinin BA was superior to other tested treatments in terms of its ability to promote germination of lentisk seeds. When tested at different concentrations, sucrose gave the best results obtained at concentrations of 1–4%, whereas high concentrations (6 and 8%) mainly decreased germination rate and there was no a regular pattern for elongation of the aerial parts of plants. With this described protocol, on average 76.67% seeds germinated 4 weeks after culture. Developed seedlings were satisfactorily acclimatized in sterilized peat, soil and perlite containing compost, with high percentage survival viability was obtained 9 months after transfer to in vivo conditions (93.33%). The results obtained showed that the enriched full-strength MS medium supplemented with 1 mg L–1 BA and 3% sucrose induced homogeneous and healthy seedling development in a period of 4 to 8 weeks of culture.