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Plant regeneration through indirect organogenesis allows obtaining genetic variability that can be used in the creation of new cultivars. The study presents a fast and effective protocol of one-step preparation of stevia (Stevia rebaudiana Bertoni) regenerants. To obtain callus tissue and shoot regeneration, leaves and nodal segments were used as primary explants, which were placed on MS (Murashige and Skoog) medium supplemented with plant growth regulators (PGRs): NAA (1-naphthaleneacetic acid – 2.0 mg·dm–3, BA (6-benzylaminopurine – 4.0 mg·dm–3), 2,4‑D (2,4-dichlorophenoxyacetic – 2.0 mg·dm–3). Callus tissue was formed on both types of explants, however, only those derived from nodal segments were proliferating. An average of 3.92 shoots per explant were obtained from leaf explants on the applied medium after 6 weeks of culture. The analysis of the morphogenetic capacity of the obtained regenerants was carried out on MS medium supplemented with PGRs – kinetin (0.25 mg·dm–3), BA (0.5 mg·dm–3). The evaluation of the mean number of shoots, mean shoot length (cm), and the mean number of nodes per shoot indicated phenotypic variability of regenerants. The use of RAPD (randomly amplified polymorphic DNA) markers confirmed the differences also at the DNA level. The proposed one-step indirect organogenesis regeneration protocol induced somaclonal variation of Stevia rebaudiana Bertoni and the obtained regenerants, after selection, could be used in the breeding of this species.

In order to improve the methods of embryo regeneration applied so far in in vitro cultures, a two-phased, innovative method was used. It involved using a liquid and solid media. Regeneration of ovules with the application of the gynogenesis technique took place mainly through the indirect organogenesis process during which callus got formed which was associated with the possibility of somaclonal variation occurrence. In order to identify the genetic variability taking plase during the culture, RAPD analyzes were performed. The DNA was isolated from the leaves of 10 regenerants, rooted and planted into the field. 5 of them were characterized by a haploid number of chromosomes whereas the others were of diploid nature. As a control, DNA originating from the leaves of a haploid plant growing in in vitro cultures on medium without growth regulators was used. Plants regenerated on media of various quantitative and qualitative composition in terms of phytohormonal characteristic. The results of the experiment showed that 9 out of 19 tested primers generate polymorphic bands while the remaining 10 amplify the same DNA fragments in all tested samples. The polymorphism effect was revealed in 4 plants with diploid genome and in 1 haploid plant. In total, 13 polymorphic bands were identified and 11 of them were detected in diploid DNA.

The present study was undertaken to evaluate efficiency of callogenesis and regeneration by somatic embryogenesis of the subendemic Iris species Iris illyrica from Croatia and to select highly regenerative donor plants/genotypes. Leaf base segments were used as explants. Callogenesis and somatic embryogenesis were induced on MS media supplemented with: (1) 4.52 micromole 2,4-dichlorophenoxyacetic acid (2,4-D) + 4.83 micromole 1-naphthaleneacetic acid (NAA) + 0.46 micromole kinetin (Kin); (2) 4.52 micromole 2,4-D + 4.6 micromole Kin; (3) 13.4 micromole NAA + 2.3 micromole Kin. Transfer of embryogenic calli onto hormone-free medium enabled the development of mature somatic embryos. Frequency of callogenesis was influenced by the donor plant. Among 15 donor plants tested, 3 of them exhibited high regeneration capability and produced 87 regenerants. Seven morphological traits were observed in order to assess phenotypic variability of flowering regenerants. Regenerants with higher values of fall and standard width and length show potential for further breeding of new varieties of Illyrian iris. Flowering and non-flowering regenerants had the same ploidy level as donor plants. Also, the nuclear DNA content of this species was estimated for the first time using flow cytometry (2C = 12.936 ± 0.038 pg).

A protocol for indirect shoot organogenesis of Solanum melongena ‘Larga Negra’ and ‘Black Beauty’ was established using hypocotyl and cotyledon derived calluses. The maximum morphogenic callus induction was observed from cultured cotyledons of 30-days old seedlings on Murashige and Skoog (MS) medium containing 2.0 mg/l α-naphthalene acetic acid and 0.5 mg/l 6-benzylaminopurine. The highest percentage of shoot regeneration and the highest mean number of shoots/callus were obtained on hormone-free MS medium. In terms of callus induction and subsequent plant regeneration, cotyledon explants were more responsive than hypocotyl explants. Regenerated shoots (2-3 cm) were rooted on MS hormone-free medium or medium containing 0.1 mg/l indole-3-butyric acid. About 90% of regenerated plantlets survived under field conditions after hardening in the glasshouse. Several somaclones exhibiting useful variation would to be proposed as initial plant material for eggplant breeding programs.