ABSTRACT
BACKGROUND: Flower color is one of the main characteristics of ornamental plants. Aurones are light yellow flavonoids produced in the petals of a limited number of plant species including snapdragon (Antirrhinum majus). As a commercially-recognized species, African violet can be found in various colors except yellow. This research, aiming at changing the petals' color of African violet from white to yellow, was conducted using the simultaneous expressions of chalcone 4'-O-glucosyltransferase (4'CGT) and aureusidin synthase (AS1) genes without the need for silencing anthocyanin biosynthesis pathway genes via both transient and stable transfer methods. RESULTS: The transient gene transfer among transgenic plants led to a clear change of petals' color from white to light yellow. This occurs while no change was observed in non-transgenic (Wild type) petals. In total, 15 positive transgenic plants, produced via stable gene transfer, were detected. Moreover, since their flower color was yellow, both genes were present. Meanwhile, the corresponding transformation yield was determined 20-30%. The transformation, expression and integration of genes among T0 transgenic plants were verified using the PCR, qRT-PCR and Southern blotting techniques, respectively. Furthermore, the probable color change of petals' cross-section and existence of Aureusidin 6-O-glucoside (AOG) compound were determined using a light microscope and HPLC-DAD-MSn analysis, correspondingly. CONCLUSIONS: Generally, the creation of aurones biosynthesis pathway is only viable through the simultaneous expression of genes which leads to color change of African violet's petal from white to yellow. This conclusion can lead to an effective strategy to produce yellow color in ornamental plant species.
ABSTRACT
Russian olive, also known as, Persian olive or oleaster (Elaeagnus angustifolia L.) is a species in the Elaeagnaceae family native to western and central Asia. In some orchards in Iran, intercropping Russian olive or Prunus trees with vegetables is a common practice. In 2018, about 130 ha of E. angustifolia orchards in Shahrood, Semnan Province, Iran showed branch wilting and dieback. Symptoms on affected trees started with yellowing of the lower leaves, followed by wilting and finally death of affected branches. Sections of stems indicated brown or black streaks in the vascular tissues under the bark. Isolations were made from discolored vascular tissues by surface-disinfesting small pieces of tissue with 0.5% sodium hypochlorite for 2 min, plating them onto potato dextrose agar amended with 25 mg/l streptomycin sulfate and incubated in the dark for 14 days at 25°C. Fungi consistently isolated from symptomatic tissues. Fungal isolates were identified as Verticillium dahliae Kleb. based on characteristics of verticillate conidiophores, hyaline, elliptical, single celled conidia measuring 4.7-6.0 × 2.3-3.4 µm (n = 100) and irregular, dark microsclerotia measuring 27-34 × 22-26 µm (n = 50) that developed after 14 days of growth at 25°C in the dark. The identification of two isolates was further confirmed by performing real-time PCR assay using a pair of specific primers for internal transcribed spacer (ITS) region of V. dahliae as previously described (Hiemstra et al. 2013). In addition, the molecular subdivision of isolates was further determined to discriminate D and ND molecular types (Keykhasaber 2017). According to molecular assays, isolates were identified as V. dahliae and grouped with ND types. The pathogenicity of isolates was evaluated by root-dipping one-year E. angustifolia seedlings (10 seedlings) into conidial suspensions of 1×107 cfu/ml. Inoculated plants were transplanted in pots containing autoclaved soil and maintained in a greenhouse at 25°C until symptoms appearance. Two seedlings were treated with sterile distilled water as controls. All inoculated seedlings started to show wilting symptoms similar to those present in naturally affected trees within 30 days after inoculation and died thereafter. Furthermore, V. dahliae was consistently isolated from symptomatic tissues. No symptoms were observed on the control plants. The pathogenicity test was repeated twice with similar results. To the best of our knowledge, this is the first report of Verticillium wilt on Russian olive trees in Iran. In Iran, Verticillium wilt is the cause of serious losses in many woody and herbaceous plants with economic importance including many trees belonging to the genus Prunus that are highly susceptible to the disease. In Shahrood (Semnan Province), most agricultural fields have a potato- or tomato- growing history. Verticillium wilt may become an important economic problem in many Russian olive and Prunus orchards in the future since their cultivation is expanding rapidly in many agricultural areas previously dedicated to tomato and potato crops, the majority of which are infested with V. dahliae. References Hiemstra, J. A., Korthals, G. W., Visser, J. H. M., Dalfsen, P. v., Sluis, B. J. v. d., and Smits, A. P. 2013. Control of Verticillium in tree nurseries through biological soil disinfestation. Pages 62-62 in: 11th International Verticillium Symposium, Georg-August-Universität,Göttingen, Germany, 5-8 May 2013, B. Koopmann and A. von Tiedemann, eds. DPG Spectrum Phytomedizin, Göttingen. Keykhasaber, M., Faino L., van den Berg, G.C.M., Hiemstra, J. A., Thomma, B. P. H. J. 2017. A robust method for discriminating defoliating and the non-defoliating pathotypes of V. dahliae. . In; Keykhasaber M. thesis 62-84. Sun, M., and Lin, Q. 2010. A revision of Elaeagnus L. (Elaeagnaceae) in mainland China. J. Systematics and Evolution 48:356-390.
