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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 expression of insecticidal genes must be induced at appropriate time and in sufficient amount to confer protection against targeted pests. However, the increased scientific reports of resistance development in insect pest against insecticidal delta-endotoxins, produced by Bacillus thuringiensis, provide impetus for the development of alternative insect management strategies. The present study was conducted to investigate the importance of targeted expression of a hybrid insecticidal gene (SN19) in potatoes. For this purpose, two plant expression vectors were constructed by cloning hybrid SN19 gene (cry1Ba-domain I–III and cry1Ia-domain II) under the control of a wound-inducible promoter isolated from Asparagus officinalis (AoPR1) and CaMV 35S promoter, and were transferred to Agrobacterium tumefaciens strain EHA 105. Four potato genotypes (Marabel, Innovator, Tokat 10/1 and Tokat 6/24) were transformed with EHA 105 strain harboring pTF101.1 35S–SN19 and pTF101.1 AoPR1–SN19 constructs. Phosphinothricin (PPT) was used at concentration of 1 mg/l for selection of primary transformants. PCR results showed the presence of both introduced SN19 and bar genes in 43 plants out of total 154 putative transgenics. Expression of SN19 protein in primary transformants was confirmed by Western blot assays. The mechanical wounding of transgenic plants exhibited more accumulated levels of SN19 proteins during post wounding period. Leaf biotoxicity assays with Colorado potato beetle (Coleoptera) and tomato leafminer (Lepidoptera) exhibited 100% mortality of the pests in primary transformants. Based on our mortality results with both constructs, we concluded that the potato transgenic lines exhibited targeted expression of insecticidal gene under the control of AoPR1 promoter upon insect wounding with eliminated toxicity of Cry protein and hence can be further used effectively in potato breeding programme. © 2017, Korean Society for Plant Biotechnology and Springer Japan KK. 111T816 Türkiye Bilimsel ve Teknolojik Araştirma Kurumu, TÜBITAK Genetic Engineering and Biotechnology Institute, Scientific and Technological Research Council of Turkey, Gebze, Kocaeli, Turkey Funding The research project was supported by a Grant from Scientific and Technological Research Council of Turkey (TÜBİTAK) Project # 111T816.

In this study, karyotype analysis of the species of Onobrychis sp. was undertaken using the squash method. The results showed that the chromosome number of Onobrychis tournefortii (Willd.) Desv., O. gracilis Besser, O. hypargyrea Boiss. is 2n = 14 and O. argyrea Boiss. subsp. argyrea Boiss., 2n = 16. The karyotype formula of O. tournefortii (Willd.) Desv. is 4m+3sm, of O. hypargyrea Boiss. and O. gracilis Besser 3m+4sm, and of O. argyrea Boiss. subsp. argyrea Boiss. 2m+ 5sm+1 st.

Erysiphe trifolii and Leveillula taurica on Onobrychis spp. in Turkey, are described and illustrated.

Amplified fragment length polymorphism (AFLP) markers were used to investigate genotypic variability among 34 alfalfa clones (M. sativa) using 15 primer combinations with restriction enzymes EcoRI, Pstl and Msel. 34 unique AFLP fragments were observed. The 15 primer pairs produced a total of 1002 fragments of which 460 were polymorphic. The number of polymorphic fragments detected per primer combination ranged 7 to 67. Furthermore, 22 clone-specific markers were also detected in the 13 clones. Data analysis was performed with NTSYSpc version 2.1 software. Genetic distance values ranged 5.9374 to 1.1453. Fifteen clones which showed the highest genetic variation were selected for producing synthetic variety of Alfalfa.

Phylogenetic relationships were studied based on morphology and nuclear ribosomal DNA internal transcript spacer (nrDNA ITS) sequences in 49 populations of 36 Onobrychis taxa belonging to 5 sections (Dendobrychis, Lophobrychis, Onobrychis, Heliobrychis and Hymenobrychis) naturally grown in Turkey. The morphological data (46 quantitative characters) were standardized and submitted for PCA analysis before the performance of hierarchical clustering analysis. In addition, a 577-bp region of the nrDNA ITS sequences was compared to generate a dendrogram using the neighbor-joining method. According to the obtained results, the Onobrychis genus is clearly divided into the Onobrychis and Sisyrosema subgenera both molecularly and morphologically; however, in terms of molecular data, while the Onobrychis subgenus showed polyphyletic features, the Sisyrosema subgenus had monophyletic features. Additionally, long-wing taxa belonging to Dendobrychis and Onobrychis and O. crista-galli belonging to Lophobrychis located in the same clade in Onobrychis showed polyphyletic features. This finding was confirmed in terms of long-wing taxa as well as morphology and indicated that these three sections were entangled with each other under the Onobrychis subgenus.

WOS:000378689600017, t: The present study addressed the successful in vitro seed germination, bulblet regeneration, increased bulblet size, hardening, and acclimatization of Persian lily (Fritillaria persica). Seed germination rate was recorded as 86.7% and 96.7% after 2 and 3 months of cold treatment at 4 °C, respectively. Bulblet explants taken from a germination experiment were cultured on Murashige and Skoog (MS) medium supplemented with 0.5–2.0 mg/L thidiazuron (TDZ). Maximum bulblet regeneration frequency and bulblets per explants were achieved on a medium containing 2.0 mg/L TDZ. Leaf disc and leaf scale explants isolated from germinated seedlings were also cultured on MS medium containing 2–6 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D) with a constant concentration of 0.2 mg/L kinetin. Both explants yielded the highest bulblet regeneration on MS medium containing 2 mg/L 2,4-D and 0.2 mg/L kinetin. A combination of 1.5 g/L agar and 1.6 g/L GELRITE resulted in a maximum bulblet diameter with a higher number of roots. The addition of 10 g/L NaCl to the culture medium resulted in the best root and leaf formation and secondary bulblet regeneration. Exposure of regenerated bulblets to 10 g/L NaCl at 4 °C for 2 months on a medium with 20 g/L sucrose was also found to be beneficial for acclimatization.

Sternbergia fischeriana is an endangered geophyte and therefore in vitro micropropagation of this plant will have great importance for germplasm conservation and commercial production. Bulb scale and immature embryo explants of S. fischeriana were cultured on different nutrient media supplemented with various concentrations of plant growth regulators. Immature embryos produced higher number of bulblets than bulb scales. Large numbers of bulblets were regenerated (over 80 bulblets/explants) from immature embryos on Murashige and Skoog (MS) medium supplemented with 4 mg l−1 6-benzylaminopurine (BA) and 0.25 mg l−1α-naphthaleneacetic (NAA) or 2 mg l−12,4-dichlorophenoxyacetic acid (2,4-D) after 14 months of culture initiation. Regenerated bulblets were kept at 5 °C for 5 weeks and then transplanted to a potting mixture.

Tobacco leaf disc explants were inoculated with Agrobacterum tumefaciens strain GV2260 carrying p35S GUS-INT to determine the influence of different co-cultivation temperatures (18 – 26 °C), periods (24 – 96 h) and media (solid and liquid) on transformation efficiency. Kanamycin-resistant shoots developed on leaf discs inoculated with Agrobacterium after 4 weeks of culture initiation. Regenerated shoots were excised and rooted in the basal medium supplemented with 100 mg dm −3 kanamycin and 250 mg dm −3 augmentin. The rooted plantlets were finally transferred to compost and confirmed by GUS assay and PCR analysis. The highest transformation frequency was achieved from the explants co-cultivated with A. tumefaciens in liquid medium for 48 h at 22 or 24 °C.

It was found that in vitro microtuberization of potato (Solanum tuberosum L.) was affected considerably by temperature and gelling agent. The best results were obtained in cultures maintained at 20–22° C in dark. Moreover, considerable increases in number, average weight and yield of microtubers was obtained on media solidified with gelrite compared to agar, when used as gelling agent. Number of microtuber formation and their weight had significant positive relationship (r = 0.630 and r = 0.771 respectively) with microtuber yield per plantlet.

The pollen size and morphology of 20 Onobrychis Adans. taxa from Turkey were studied using both light and scanning electron microscopy. Of the taxa studied, Onobrychis fallax Freyn & Sint. and Onobrychis elata Boiss. & Balansa showed the smallest size of pollen grains, while the largest were found in Onobrychis aequidentata (Sibth. & Sm.) d'Urv. and Onobrychis viciifolia Scop. The pollens of all taxa studied represent the tricolpate type of pollen and are prolate in shape. The type of exine found in the studied taxa is reticulate (sect. Dendobrychis DC. and sect. Laphobrychis Hand.-Mazz.), suprareticulate (sect. Onobrychis), or microreticulate (sect. Hymenobrychis DC. and sect. Heliobrychis Bunge). A correlation is observed between the pollen size and chromosome number.

Efficient and reproducible shoot regeneration has been established from immature cotyledons and embryo axes of Hungarian vetch. Immature embryo axes showed higher regeneration capacity than immature cotyledons in most media tested. Immature cotyledons appeared to produce the highest frequency of shoot regeneration on 20 μM 6-benzylaminopurine (BA) and 2.5 μM α-naphthaleneacetic acid (NAA)-containing medium; whereas, shoot regeneration on immature embryo axes was best achieved on a medium supplemented with 5 μM each of BA and NAA. Regenerated shoots were excised and rooted in half-strength MS medium supplemented with 5 μM indole-3-butyric acid (IBA). Rooted plantlets were acclimatized to ambient conditions and later established under greenhouse conditions.

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.

An efficient in vitro bulblet production procedure from immature zygotic embryos of endemic and endangered Muscari muscarimi Medik. was described in the current study. Zygotic embryos were first isolated from immature seeds and cultured on different nutrient media compositions supplemented with various combinations of ?-naphthalene acetic acid (NAA), picloram, dicamba, 6-benzylaminopurine (BAP), and thidiazuron (TDZ). The best bulblet regeneration (59 bulblets per explant) was achieved in Murashige and Skoog (MS) medium containing 4 mg/L BAP and 0.5 mg/L NAA after 1 year of culture initiation. Regenerated bulblets were then transferred into MS medium without plant growth regulators for rooting. Bulblets produced well-developed root systems and increased their size on this medium after 2 months. All rooted bulblets were successfully transplanted into a potting mixture and acclimatized to ambient conditions. © TÜBİTAK.

Seven genotypes of winter durum wheat (Triticum durum Desf.) were cultured to establish an efficient method of callus formation and plant regeneration from mature embryo culture, and to compare the responses of immature and mature embryo cultures. Immature embryos were aseptically dissected from seeds and placed, with the scutellum upwards, in dishes containing Murashige and Skoog's (MS) mineral salts and 2mg 2,4- dichlorophenoxyacetic acid (2,4-D) per litre. Calli and regenerated plants were maintained on 2,4-D-free medium. Mature embryos were moved slightly on the imbibed seeds. For callus formation, the seeds with moved embryos were placed, furrow downwards, in dishes containing 8 mg 2,4-D per litre. The developed calli and regenerated plants were maintained on the MS medium. Plants regenerated from both embryo cultures were vernalized and grown to maturity in soil. Variability was observed among the wheat genotypes tested for various culture responses in both explant cultures. Callus induction rate and regeneration capacity of callus were independent of each other. Mature embryos have a low frequency of callus induction but a high regeneration capacity. Considering availability, rapidity and reliability, this form of mature embryo culture can be used as an alternative method for immature embryo culture.

Immature 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.

Transformation of fenugreek ( Trigonella foenumgraecum) was carried out with A281 oncogenic strain of Agrobacterium tumefaciens using root, cotyledon and hypocotyl explants excised from 1-week-old seedlings, which showed that the plant was highly susceptible to transformation. Tumors ( calli) were selected on 50 mg dm(-3) kanamycin. They were analyzed for beta-glucuronidase (GUS) expression. Presence of uidA (gus) gene, was confirmed by polymerase chain reaction (PCR) amplification.

Efficient and reproducible shoot regeneration has been established from immature zygotic embryos of lesser burnet on MS medium containing 4 μM BAP and 10 μM NAA. Regenerated shoots were best rooted in half-strength MS medium supplemented with 10 μM NAA and later established in the greenhouse.