Murat Özgen

Efficient callus induction and plant regeneration from mature embryo culture of winter wheat (Triticum aestivum L. ) genotypes

Abstract Immature and mature embryos of 12 common winter wheat (Triticum aestivum) genotypes were cultured in vitro to develop an efficient method of callus formation and plant regeneration from mature embryo culture, and to compare the responses of both embryo cultures. Fifteen days after anthesis, immature embryos were aseptically dissected from seeds and placed with the scutellum upwards on a solid agar medium containing the inorganic components of Murashige and Skoog (MS) and 2 mg/l 2,4-dichlorophenoxyacetic acid (2,4-D). Mature embryos were moved slightly in the imbibed seeds. The seeds with moved embryos were placed furrow downwards in dishes containing 8 mg/l 2,4-D for callus induction. The developed calli and regenerated plants were maintained on 2,4-D-free MS medium. Plants regenerated from both embryo cultures were vernalized and grown to maturity in soil. Regenerated plantlets all maintained the hexaploid chromosome number. A strong genotypic effect on the culture responses was found for both explant cultures. Callus induction rate, regeneration capacity of callus and number of plants regenerated were independent of each other. Mature embryos had a high frequency of callus induction and regeneration capacity, and therefore, being available throughout the year, can be used as an effective explant source in wheat tissue culture.

The effect of a submersion pretreatment on in vitro explant growth and shoot regeneration from hypocotyls of flax (Linum usitatissimum)

This study was carried out to determine the effect of temporary submersion of hypocotyl segments in water on in vitro explant growth and shoot regeneration on MS (Murashige and Skoog, 1962) medium supplemented with 1 mg l−1 BAP (6-benzylaminopurine) and 0.02 mg l−1 NAA (naphthaleneacetic acid) in three flax cultivars. It was observed that water-treated hypocotyl explants gave rise to the highest values with respect to shoot regeneration percentage, shoot number per hypocotyl, shoot length and total shoot number per Petri dish, successful rooting and plantlet establishment. This procedure may be applicable for other species cultured in vitro.

Prolific shoot regeneration from immature embryo explants of sainfoin (Onobrychis viciifolia Scop.)

SummaryImmature cotyledons and embryo axes of sainfoin were cultured on Murashige and Skoog (MS) media supplemented with various concentrations of 6-benzylaminopurine (BAP) and a-naphthaleneacetic acid (NAA) to induce adventitious shoot regeneration. The highest frequency of shoot regeneration occurred following an initial callus growth on a MS medium containing 0.5 mg/l BAP and 2 mg/l NAA. Immature embryo axes showed higher regeneration capacity than immature cotyledons, however, shoot elongation was best achieved on immature cotyledons. Regenerated shoots were excised and rooted in half strength MS medium with 1 mg/l indole-butyric acid (IBA) or 1 mg/l NAA. The rooted plantlets were finally transferred to compost.

High frequency adventitious shoot regeneration in sainfoin

A procedure is described for the rapid and efficient adventitious shoot regeneration from leaflets, petioles and stems of field-grown sainfoin plants. All explants formed shoots on a range of media supplemented with 6-benzyladenine (BA) and α-naphthaleneacetic acid (NAA). Stem explants appeared to have better regeneration capacity than leaflet and petiole explants in most media tested. The highest frequency of shoot regeneration was achieved from stem segments on a medium containing 20 μM BA and 0.5 μM NAA. Regenerated shoots rooted in half-strength Murashige and Skoog medium containing 5 μM indole-3-butyric acid and later established well under greenhouse conditions.

Cytoplasmic effects on the tissue culture response of callus from winter wheat mature embryos

The cytoplasmic effects on embryo culture responses were studied by using calli from mature embryos of four genotypes of winter wheat (Triticum aestivum L.). Significant reciprocal differences were found for tissue culture response of callus, indicating that a cytoplasmic effect may be involved. In general, cytoplasm positively affected callus induction, regeneration capacity of callus, culture efficiency and numbers of regenerated plants. The nature of the effect of the cytoplasm is shown to depend on the genotype. The potential for improving the responses of tissue culture by cytoplasmic changes was observed.

The effect of hybrid vigor on callus induction and plant regeneration from mature embryo culture of barley (Hordeum vulgare)

The influence of hybrid vigor on callus induction, callus weight, regeneration capacity of callus, culture efficiency and number of regenerated plants from mature embryo cultures of winter barley (Hordeum vulgare L.) hybrid was studied. A total of 15 F1 hybrids and their parents were used for mature embryo culture. The statistical analysis of the results revealed that positive heterosis was noticed for all these characters except for callus weight. Plants regenerated in vitro were successfully established in soil.

The Effect of Seed Size on Tissue Culture Response of Callus from Endosperm-supported Mature Embryos of Barley (Hordeum vulgare L.)

Although immature embryos are the most suitable explants to regenerate cereals, their limited availability throughout the year hampers frequent experimentation. In order to circumvent this problem, endosperm-supported mature embryos, which are available throughout the year, are proposed for utilization in cereal tissue culture studies. In the present study, the effect of seed size on in vitro seed germination, seedling growth, callus induction and plant regeneration, as well as the relationships between these parameters, were investigated in three barley (Hordeum vulgare L.) genotypes. Seeds were designated as large or small for each genotype and seed germinations were achieved in Petri dishes between two sheets of pre-wetted filter paper. Germination percentages of seeds, seedling growth rates, shoot and root lengths, and seedling fresh and dry weights were examined. Mature embryos from imbibed and dehulled seeds were aseptically moved slightly with a scalpel. For callus induction, 8 mg 1−1, 2,4-D dissolved in water, and for plant regeneration, hormone-free MS medium, was utilized. Highest values with respect to seed germination percentage, seedling growth, callus induction and plant regeneration were obtained using large seeds. Rooting of regenerated shoots and plantlet recovery were also successfully achieved. Based on the significant positive correlations observed between all parameters, we concluded that cultures derived from endosperm-supported mature embryos within large seeds should be employed for achieving superior tissue culture response.

Phylogenetic relationships among Triticum L. and Aegilops L. species as genome progenitors of bread wheat based on sequence diversity in trnT-F region of chloroplast DNA

Cultivated wheat, (Triticum aestivum L.), is one of the most important food crops in the world. The Aegilops L. genus is frequently utilized by plant breeders for improving the current wheat cultivars due to their close relationships. Therefore, understanding the phylogenetic relationships among the species of these genera is not only valuable for plant taxonomy, but also for plant breeding efforts. The presented phylogenetic analysis was based on the sequences of trnT-F chloroplast DNA containing three non-coding sub-regions. Twelve genotypes belonging to four species of Triticum L. genus and twenty-four genotypes belonging to eight species of Aegilops genus were used in the current study. The results postulated a close genetic relationship between diploid Aegilops species containing the BB genome and polyploid Triticum species. With the exception of Aegilops cylindrica Host (CCDD), all other Aegilops species having the CC genome were alienated from Aegilops speltoides Tausch (BB) and clustered together. These two clusters joined by a third cluster including the AA genome containing diploid Triticum species.

Callus Formation and Plant Regeneration from Cultured Embryos of Diploid and Tetraploid Winter Ryes (Secale cereale L)

The ability of immature, mature and endosperm-supported mature embryos of diploid and tetraploid winter ryes (Secale cereale L) was tested to compare the callus induction and plant regeneration. Immature embryos were obtained from field grown rye. Immature embryos were aseptically excised and placed, with the scutellum upwards, on the callus culture medium consisted of Murashige and Skoog (MS) mineral salts supplemented with 2 mg l−1 2,4-dichlorophenoxyacetic acid (2,4-D). Mature embryos were aseptically excised the imbibed seeds and placed, scutellum up, on MS medium supplement with 2 mg l−1 2,4-D. Endosperm-supported mature embryos were moved slightly (not set free) in the imbibed mature seeds. The seeds with moved embryos were placed furrow downwards in dishes containing 8 mg l−1 2,4-D for callus induction. The developed calli and regenerated plant were maintained on hormone free MS medium. Comparison of the responses of the three explants used indicated that endosperm-supported mature embryo was the most useful explant for plant regeneration in both diploid and tetraplold ryes. This is the first report of winter ryes plants having been regenerated from endosperm-supported mature embryos.