Identification of C-banded chromosomes in meiosis and the analysis of nucleolar activity in Avena byzantina C. Koch cv ‘Kanota’

Summary The Giemsa C-banding technique was used to identify individual meiotic and somatic chromosomes in 21 monosomic lines of Avena byzantina C. Koch cv Kanota (genome designation AACCDD). The hexaploid complement is composed of three sets of seven chromosome pairs. The heterochromatin in the putative diploid progenitors is located at the telomeres (genome A), at the centromeric and interstitial regions (genome C), or more evenly spread throughout the set (genome D). Comparisons based on C-banding between A. byzantina and its diploid progenitor species allowed us to allocate individual chromosomes into specific genomes. The C-banding technique may be useful for interspecific chromosome pairing analyses. Nucleolar activity and competition were studied using a silver-staining procedure. Only three chromosome pairs showed nucleolar organizer regions, thus indicating that nucleolar competition occurs naturally in hexaploid oats.     

Micropropagation of Achillea filipendulina cv. ‘Parker’

Achillea filipendulina (family Asteraceae) is widespread throughout temperate North America. In order to clean stock plants from endemic fungal and bacterial contaminations a method for large-scale propagation of A. filipendulina through meristem culture was sought and found and is described in this paper. The best conditions for propagating A. filipendulina was found to be MS (Murashige and Skoog) salt medium supplemented with 3% sucrose and 1 mg l^–1 IAA (indole-3-acetic acid) plus 2 mg l^–1 BA (6-benzyladenine) under 16 h of cool fluorescent light. Rooted plants were successfully acclimatized within a short time after propagating on this medium. The propagation via tissue culture did not affect plant's presentation. The use of clean stock plants made it possible to increased Israeli production of Achillea from about 150,000 stems a year to about 1,300,000 stems a year.     

Measurement of selected trace elements in Olea europaea L. cv. ‘Sigoise’

BackgroundOlive-trees (Olea europaea L.) are the dominant rustic trees cultivated in the Mediterranean agricultural zones. Major and micronutrients play an indispensable role in their plant physiological functions although; the effect of trace elements on metabolic processes has not been sufficiently investigated, especially in olive-trees.MethodsIn the current study, we have used X-ray fluorescence (XRF) spectrometry to determine selected major and trace elements (Br, Cu, Fe, K, Mn, P, Rb and Zn) in the main olive cultivar cultivated in Algeria, cv.‘Sigoise’. Certified reference materials viz. IAEA-336 (Lichen) and NIST-1646a (Estuarine sediment) were evaluated simultaneously with the soil and plant samples for quality control of the analytical method.ResultsThe results show that Fe and Mn concentrations were superior in leaves than fruits. However large amounts of K, Cu and Rb were accumulated in the olive-fruits. The contents of all chemical elements were above the threshold limits for possible plant nutrient deficiencies, except for P whose concentration was in borderline requirement of olive trees. High values of a translocation factor index were found for K, Cu and Rb (TFs > 4). Principal component analysis (PCA) indicated that K was highly related with olives-fruits, suggesting that the fruit was the principal organ of K storage. Furthermore, dietary element intake through consuming olives was also estimated and compared to recommended daily intakes (RDIs) and daily permissible limits (DPLs). The estimations of chemical element intakes were below the DPLs set by WHO/FAO guidelines for human nutrition.ConclusionThe present work indicates that the concentrations of macro- and microelements (Cu, Fe, K, Mn and Zn) were above the threshold limits for possible plant deficiencies except for P, and this cultivar can easily accumulate high amount of K in their organs (predominance in olives). These findings will be used to achieve efficient fertilization for O. europaea orchards.     

Micropropagation of Calluna vulgaris cv. ‘H.E. Beale’

Axillary shoot producing cultures were obtained from microcuttings and shoot tips of Calluna vulgaris cv. H.E. Beale. For cultures derived from microcuttings the highest multiplication rate of 38 shoots (5 mm or longer) was obtained on a reduced salt medium with the addition of 0.5 mgl^-1 2-isopentenyladenine (2iP) during an 8 week subculture. For shoot tip derived cultures 0.2 mgl^-1 6-benzyladenine (BA) was the best cytokinin and led to a multiplication rate of 26 for a 6 week subculture. The addition of 1 g/l casein hydrolysate to a multiplication medium enhanced shoot proliferation in presence of 0.5 mgl^-1 BA.Despite various auxin treatments shoots formed no roots in vitro but rooted readily if transferred to a peat substrate ex vitro. A high rooting percentage (80%) was also obtained with shoots taken from the end of a multiplication phase and rooted directly. An additional subculture on low auxin containing media before transfer to peat substrate is recommended because the shoot condition can be improved in this way. A high number of rooted plantlets was produced, so the methods described will allow mass propagation.     

Gametoclonal variation detected in the nuclear ribosomal DNA from doubled haploid lines of a spring wheat (Triticum aestivum L., cv. ‘César’)

Summary The organization of the nuclear ribosomal DNA from a parental line of wheat (Triticum aestivum L., cv. César) and its anther-derived first cycle and second cycle doubled haploid lines has been analyzed by DNA-DNA molecular hybridization. Restricted DNA has been probed by three subclones of wheat nuclear ribosomal DNA covering the entire repeat unit. No significant difference was detected in the extent of methylation of ribosomal DNA of the doubled haploid lines with respect to the parental line. On the other hand, a variation has been found in the organization of the nontranscribed spacer region of ribosomal DNA of the first cycle doubled haploid line. This variation remains stable after a second cycle of in vitro androgenesis. However, one out of five second cycle doubled haploid lines so far tested showed an additional hybridization band present in the parental line but lacking in the first cycle doubled haploid line.     

Somatic embryogenesis in Rosa chinensis cv. ‘Old Blush’

Plant Cell, Tissue and Organ Culture (PCTOC) - Rose is one of the most widely used ornamental flowers in the world. However, the low induction rates for most rose somatic embryos means that it is...     

Culture-independent analysis of an endophytic core microbiome in two species of wheat: Triticum aestivum L. (cv. ‘Hondia’) and the first report of microbiota in Triticum spelta L. (cv. ‘Rokosz’)

The main goal of the study was to determine the structure of endophytic bacteria inhabiting different parts (endosperm, germ, roots, coleoptiles, and leaves) of two wheat species, Triticum aestivum L. (cv. ‘Hondia’) and Triticum spelta L. (cv. ‘Rokosz’), in order to provide new knowledge about the stability and/or changeability of the core microbiome in different plant organs. The endophytic core microbiome is associated with plants throughout their whole life cycle; however, plant organs can determine the actual endophytic community. Therefore, next generation sequencing with MiSeq Illumina technology was applied to identify the endophytic microbiome of T. aestivum and T. spelta. Bioinformatic analyses were performed with the use of the DADA2(1.8) package and R software (3.5.1).It was demonstrated that wheat, which is an important crop plant, was associated with beneficial endophytic bacteria inside the endosperms, germs, roots, leaves, and coleoptiles. Importantly, for the first time, biodiversity was recognized in the coleoptiles of the investigated wheat species. Flavobacterium, Pseudomonas and Janthinobacterium were shown to be common genera for both tested wheat cultivars. Among them, Pseudomonas was found to be the only endophytic genus accompanying both wheat species from the endosperm stage to the development of the leaf. Paenibacillus was recognized as a core genus for the ‘Hondia’ cv., whereas Pedobacter and Duganella constituted the core microbiome in the ‘Rokosz’ cv. In addition, the first insight into the unique and yet unrecognized endophytic microbiome of T. spelta is presented.     

Agrobacterium-mediated transformation of Solanum tuberosum L. cv. ‘Russet Burbank’

Stem sections from shoot cultures maintained in vitro were used to produce transgenic plants of the potato, Solanum tuberosum L. cv. Russet Burbank. Stem internode pieces inoculated with Agrobacterium tumefaciens containing coat protein genes from potato virus X and potato virus Y, produced shoots with a frequency of 60% in the absence of selection and 10% on medium containing 100 mg/l kanamycin monosulfate. Regenerated shoots were assayed for kanamycin resistance by placing stem segments on callus induction medium containing an increased level of kanamycin. Of a total 255 regenerated shoots, 47 (18%) were kanamycin resistant. Of the kanamycin resistant shoots, 25 (53%) expressed the PVX or PVY coat protein genes as assayed by enzyme-linked immunosorbent assay or Western immunoblot analysis.     

Effect of the rootstock on the occurrence of lime-induced chlorosis of potted Vitis vinifera L. cv. ‘Pinot blanc’

The chlorosis susceptible Vitis vinifera L. cv. Pinot blanc was grafted on two hybrid rootstocks with different iron efficiency, as follows: V. Berlandieri × V. rupestris 140 Ru (iron-efficient) and V. riparia × V. rupestris 101-14 (iron-inefficient). The grafted vines were grown in pots of a calcareous and a non-calcareous soil. The shoot growth was periodically checked and leaves, selected at two different times (at the middle of the annual growing period), were assayed for total chlorophyll, ferrous iron, ash alkalinity, percentage of dry matter and chlorosis score. At the end of the growing cycle the roots were oven-dried and weighed. The most significant findings of the trial were: (a) the soil strongly affected the shoot growth, with canes about twice as long in the non-calcareous soil; (b) the iron-efficient rootstock (140 Ru) did not induce chlorosis when growing on the calcareous soil, while the opposite occurred with the iron-inefficient rootstock (101=14); and (c) a high ash alkalinity occurred in light chlorotic leaves compared to green ones, under the same iron concentration.     

Effect of potassium humate on apple cv. ‘Golden Delicious’ cultured in vitro

The effects of humic substances on in vitro culture of Golden Delicious apple are reported. Potassium humate (KH) when used in proliferation showed a negative interaction with BA while it enhanced rooting when IBA was not present in the culture medium. In the presence of IBA, KH increased root number and reduced root growth. The highest concentration tested, 500 mg l^-1, caused a drastic reduction in root system development. 50 mg l^-1 KH hastened rooting and plants grew more rapidly when transferred to soil.     

A stable reddish purple anthocyanin in the leaf ofGynura aurantiaca cv. ‘Purple Passion’

The anthocyanin (GAA) in the epidermis and hair of the leaf ofGynura aurantiaca cv. ‘Purple Passion’ was isolated and identified as cyanidin tetra-glucoside acylated by three molecules of caffeic acid and one molecule of malonic acid. GAA was also isolated from the lower epidermis of the leaf ofG. bicolor DC. GAA showed a very stable reddish purple color from weakly acid to neutral pH region, but the color of the deacylated compound disappeared rapidly in the same region. This indicated that the attached organic acids must play an essential role in the stabilization of the color. Comparison of the profiles of the visible absorption spectra of the intact epidermal peels and cells ofG. aurantiaca andG. bicolor with those of GAA dissolved in various pH solutions suggested that the pH of the epidermal vacuole containing GAA was nearly 4.3. GAA was indistinguishable from the anthocyanin (rubrocinerarin) which we had previously isolated from the purplish red flowers ofSenecio cruentus DC. by means of UV-Vis, NMR and Mass spectra. Deceased     

Analysis of capsaicinoid biosynthesis pathway genes expression in callus cultures of Capsicum chinense Jacq. cv. ‘Umorok’

The placental tissue of the highly pungent chilli cultivar, Capsicum chinense Jacq. cv. ‘Umorok’, is used as explants for callus induction. Callus cultures were subcultured after every 32 days and growth curves for a period of six consecutive growth cycles were studied till a stable capsaicinoids producing callus cultures were obtained. The capsaicinoids content in placental tissue explants decreased gradually during the first 2 months of culture as the explants dedifferentiated to form friable callus while the biomass and capsaicinoid content did not show much change in the subsequent growth cycles. The maximum callus biomass of 7.8 g freshweight (FW) or 0.56 g dry weight (DW) per culture were obtained on the 24th day of every growth cycle and the maximum average capsaicinoids content (1.6 mg g^?1 FW capsaicin and 0.78 mg g^?1 FW dihydrocapsaicin) were obtained on the 20th day of every growth cycle. To investigate the underlying dynamics for capsaicinoid biosynthesis during callus formation, comparative gene expression analysis of the genes involved in capsaicinoid biosynthesis pathway were also studied by qRT-PCR analysis. When compared with placental tissue, all the studied genes showed reduced expression during callus formation, especially putative aminotransferase (pAMT) and pungent gene 1 (Pun1), which were extensively down regulated from the 3rd month onwards in the callus cultures. Therefore, the present study revealed that the down-regulated expression of mainly two putative genes in capsaicinoid biosynthetic pathway (pAMT and Pun1) resulted in lower accumulation of capsaicinoids in callus cultures compared to placental tissues of fruits.

     

Influence of some herbicides on the reproduction of tobacco mosaic virus inNicotiana tabacum cv. ‘Samsun’

The application of 10^?3 g 1^?1 of 2,4 dichlorophenoxyacetic acid, 2-methyl-4-chlorophenoxyacetic acid, 3-amino-1,2,4-triazole and 6-chloro-2-ethylamino-4-isopropyl-amino-1,3,5-triazine toNicotiana tabacum cv. ‘Samsun’ plants inoculated with tobacco mosaic virus results in an increase in the content of this virus in the tissues. When whole plants are used, TMV content increases by 20% after herbicide application; when leaf dises are used, the amount of the virus can be doubled by the herbicide in comparison with control untreated inoculated discs. The results clearly show that the used non-selective herbicides act as stimulators of virus biosynthesis, probably via enhanced pentose phosphate cycle activity which enables an enhanced formation of viral RNA.     

Induction of Haploid Morphogenic Calluses from in vitro Cultured Anthers of Prunus Armeniaca cv. ‘Harcot’

Apricot (Prunus armeniaca) ‘Harcot’ anthers, were cultured in vitro for the production of haploid plants. The best androgenic response was achieved with Nitsch and Nitsch (1969) medium, supplemented with 4.52 μM 2,4-D, 4.52 μM zeatin, 2.85 μM IAA and 40 g l⁻¹ sucrose. Cultures were maintained in the dark for 8 days, at 28°C, followed by transfer to a 16-h photoperiod, with 35 μm m⁻² s⁻¹ light intensity and 24/22°C day/night temperature. The androgenic response was correlated with the floral bud size, its phenologic stage and the level of microspore evolution. Anthers containing microspores at the tetrad/uninucleate stage were the most appropriate. The ploidy level of the calluses was evaluated by flow cytometry revealing that they range from haploid to octaploid. Mixoploid calluses have also been identified. Histological studies showed that the haploid calluses have their origin in the microspores. Nodular structures consisting of cells with dense cytoplasm and differentiated xylem elements were observed and were surrounded by an autofluorescent layer, probably due to cutin deposition.     

Interactions of calcium,nitrogen and potassium with phosphorus on the symptoms of toxicity in Grevillea cv. ‘Poorinda Firebird’

In two hydroponic experiments it was found that phosphorus induced necrotic leaf symptoms and reduced the growth of Grevillea cv. Poorinda Firebird, a member of the Proteaceae family in which a number of species are known to be affected by phosphorus toxicity. The addition of high levels of calcium was found to aggravate these effects while high nitrogen or potassium levels alleviated them. Tissue analysis of plants receiving high levels of phosphorus showed a tenfold increase in leaf phosphorus compared with plants receiving no phosphorus. Lesser increases were measured in root and stem tissues.     

Cytogenetic events in the endosperm of amphiploid Avena magna × A. longiglumis

Journal of Plant Research - This study analysed cytogenetic events occurring in the syncytial endosperm of the Avena magna H. C. Murphy & Terrell × Avena longiglumis Durieu...     

In vitro regeneration of Alstroemeria cv. ‘Yellow King’ by direct organogenesis

The common techniques for the in vitro production of Alstroemeria plants are based on rhizomes as explants, which have low multiplication rates and a high risk of carrying viral diseases. To overcome these problems, we developed a protocol for the in vitro regeneration of Alstroemeria cv.‘Yellow King’, by testing for shoot induction several explant sources (leaf, stem apices, rhizomes and immature inflorescence apices), temperature and light/dark regimes, hormone and salt concentrations. For shoot multiplication and rooting, several hormone concentrations were tested. We found that only the young floral apices produced adventitious shoots by direct organogenesis. The highest shoot induction rate (10.4 shoots per explant) was obtained by incubation in the dark for 15 days at 8 °C followed by 15 days at 25 °C and a 16-h/8-h light/dark regime, on a Murashige and Skoog (1962) liquid medium at 50% of the salt concentration, supplemented with 2.5 mg l⁻¹ KIN, 1.5 mg l⁻¹ BA and 1.0 mg l⁻¹ NAA, using a piece filter paper to support the explant. The highest shoot multiplication rate (9 shoots per explant) was obtained on a liquid MS medium at full strength supplemented only with BA at 1.0 mg l⁻¹. In vitro rooting of shoots was induced also on a liquid MS medium, either with or without plant hormones.     

Plant regeneration via somatic embryogenesis of Elymus sibiricus cv. ‘chuancao No. 2’

The mature seeds, mesocotyls, and young leaf tips of Elymus sibiricus L. cv. ‘chuancao No. 2’ were cultured on Murashige and Skoog (MS) medium supplemented with 5.0 mg/L 2,4-dichlorophenoxyacetic acid (2,4-d) and 0.05 mg/L kinetin in the dark at 26°C, the calluses were produced. The rate of callus regeneration depended on the explants source and plant growth regulators. Plants regenerated from whitish-yellow-coloured compact nodular callus formed after subculturing for 8 weeks. Higher frequency (54%) of shoot differentiation was obtained from the embryo tissues of mature seed than from either mesocotyls (24%) or young leaf tip tissues (6%) when these calluses from different types of explants were cultured on plant regeneration medium containing half strength MS salts supplemented with 0.1 mg/L kinetin, 1.5 mg/L 2,4-D and 20 g/L sucrose. The green plants were rooted within 6 weeks in the root regeneration medium, and over 97% of these soil-established plants were obtained in the greenhouse when potted in a sand and peat mixture medium.