In vitro Regeneration of Japanese Burdock (Arctium lappa L.) from Cotyledon and Leaf Derived Explants

Considering the vegetable and medicinal values, a micropropagation protocol has been established for Japanese Burdock (Arctium lappa L.) by culturing the explants of cotyledon and leaf obtained from in vitro grown seedlings. Direct shoot regeneration was achieved from cotyledon and leaf explants on MS fortified with 4.0 μM BAP and 2.0 μM IBA or NAA after 5 weeks of culture. In addition, both the explants also formed callus from their cut margins within 6 weeks of cultivation on medium complemented with 6.0 μM BAP and 4.0 μM IBA or NAA. Adventitious shoots were also redeveloped through indirect organogenesis from the cotyledon and leaf-derived callus within 10 weeks of culture on MS containing 4.0 μM BAP and 2.0 μM IBA or NAA. The highest rate of shoot reproduction was attained at the third subculture, and more than 12.6 shoots were formed per callus clump. Within 4 weeks of transfer to the rooting medium on MS containing 6.0 μM IBA, the cultured micro-shoots produced highest 5.3 roots per cultured shoot. Rooted plantlets were successfully established on a soil-composed-sand mixture under natural condition with 93.3% survival rate Plant Tissue Cult. & Biotech. 31(2): 123-134, 2021 (December)

Establishment of Tissue Culture Regeneration System for Medicago ruthenica L. cv. ‘Zhilixing’

Background: Medicago ruthenica L. ‘Zhilixing’ is a new variety with superior forage and seed yield compared to the wild type. The cold, drought and salt tolerance of Zhlixing are better than those of alfalfa, suggesting that this variety can serve as a high-quality genetic resource for improving the stress resistance of alfalfa. However, because of the lack of tissue culture regeneration system, it is difficult to perform genetic transformation studies on stress resistance genes. This study aimed to establish an efficient tissue culture regeneration system for Zhilixing variety. Methods: Three types of explants were selected and tested on four types of basal media supplemented with different combinations of auxin and cytokinin for callus induction and differentiation, based on orthogonal tests to select the combinations of auxin and cytokinin suitable for callus induction and differentiation. Two-factor combination method was used to formulate a suitable rooting medium. Result: The hypocotyledonary axis was found to be an excellent explant for callus induction on MS medium. The optimum callus induction medium contained thidiazuron (TDZ, 0.5 mg/L), 2,4-dichlorophenoxyacetic acid (2,4-D, 1.0 mg/L) and naphthaleneacetic acid (NAA, 0.5 mg/L) where the callus induction rate was 93.33%. The differentiation medium was supplemented with TDZ (0.75 mg/L), 2,4-D (0.25 mg/L) and 6-benzyladenine (6-BA, 1.5 mg/L) where the differentiation rate was 63.33%. Thidiazuron played the key role in both processes of callus induction and differentiation. Half-strength MS containing 0.1 mg/L of NAA was the most efficient rooting medium.

IN VITRO PROPAGATION OF PLUM ROOTSTOCK DOCERA 6

This paper describes research on the application of tissue culture techniques to the micropropagation of interspecific rootstock ʹDocera 6ʹ. The experimental work was carried out in the period 2017-2018, in the in vitro propagation laboratory of the Fruit Growing Institute Plovdiv. Аxillary buds were employed as initial explantеs in two different seasons (March-May; September-October). The action of the mineral medium was studied in the multiplication stage. The best result was obtained on LS medium included BAP 0.5 mg/l and IAA 0.05 mg/l. Тhe obtained average multiplication rate is 3.08. The concentration of auxin applied to the basal medium influence the quality of the root system Treatment with high concentrations of IBA added to the rooting medium gives the best results (V5). Тhe influence of the season on growth and development of micropropagated ʹDocera 6ʹ rootstock during ex vitro acclimatization is also part of our research. The spring acclimatization gives better results than the autumn.

The effects of different factors on propagation by hardwood cuttings of some coniferous ornamental plants

Chamaecyparis lawsoniana ‘Ellwoodii’, Cryptomeria japonica ‘Elegans’ and x Cupressocyparis leylandii have great importance in terms of usage areas as ornamental plant. The overcoming the problem, that may be encountered in generative propagation of these taxa, using vegetative propagation method are very important for producers dealing with park and garden works. It was investigated that the effects of different greenhouse medium, rooting medium and phytohormones on propagation by hardwood cutting of studied taxa. In this study, three greenhouse medium (Greenhouse-1, 2 and 3 medium), two rooting medium (perlite and peat) and four phytohormones (IBA 1000, IBA 5000, NAA 1000 and NAA 5000 ppm) were selected. The first root and callus formation dates, rooting percentage, callus percentage, root length and the number of roots were determined. The results showed that the highest rooting percentages for C. lawsoniana ‘Ellwoodii’ were obtained as 100% in peat rooting medium (IBA 1000 ppm) at Greenhouse-2 medium and Greenhouse-3 medium and perlite rooting medium (control) at Greenhouse-3 medium. On the other hand, the highest rooting percentages for C. japonica ‘Elegans’ were obtained as 100% in IBA 1000, IBA 5000 and NAA 1000 ppm treatments at Greenhouse-1 medium, while this value for x C. leylandii occurred in NAA 5000 ppm treatment (73.33%) at Greenhouse-2 medium, in perlite rooting medium for both. The conditions required for the best rooting in cutting propagation vary according to the species studied, and generally, auxin applications, rooting medium temperature 5 °C higher than air temperature and use of perlite rooting medium can be recommended.

Shoot Proliferation, Callus Induction And Plant Regeneration In Tripsacum Laxum Nash

The peduncles of Tripsacum laxum Nash were used as explants to induce axillary shoots. Multiple shoots were proliferated on Murashige and Skoog (MS) medium to establish, for the first time, efficient shoot proliferation and plant in vitro regeneration systems. Optimal shoot proliferation medium was MS with 3.0 mg/L 6-benzyladenine (BA) and 0.2 mg/L α-naphthaleneacetic acid (NAA), resulting in a shoot proliferation coefficient of 11.0 within 45 d. Optimal rooting medium was MS with 0.1 mg/L NAA and/or 0.1 mg/L indole-3-butyric acid (IBA), inducing 100% root formation from shoots within 30 d. When young roots, leaf sheaths and shoot bases were used as explants, MS medium with 1.0 mg/L thidiazuron (TDZ) and 0.2 mg/L BA induced most shoots, with the least callus. Shoot bases induced beige-white callus and shoots directly on MS medium with 1.0 mg/L TDZ and 0.2 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D), while leaf sheaths induced beige-white callus and shoots directly on MS medium with 1.0 mg/L TDZ and 0.2 mg/L BA. Rooted plantlets showed 99.3% survival when transplanted into a substrate of vermiculite: peat soil (1:3, v/v).

Micropropagation and experimental field cultivation of Pulsatilla turczaninovii Kryl. et Serg. (Ranunculaceae)

Pulsatilla turczaninovii is an important medicinal plant, valued for high ornamental value of melliferous flowers. We assessed the efficiency of reproduction under in vitro conditions and the ex situ growth capacity of this important representative of the world flora. The seed germination percentage was assessed, followed by determination of micropropagation rate and rooting efficiency. Then, the possibility of plant development in three consecutive growing seasons was assessed. The in vitro germination percentage was approximately 55%. The highest multiplication coefficient, amounting to 5.17, was obtained on modified MS medium supplemented with 2.5 mg L−1 2iP and 1.0 mg L−1 IAA. Our study provided unique insight on biochemical background of root regeneration in P. turczaninovii. In comparison with standard auxin-supplemented rooting medium, the treatment with 1.0 mg L−1 level of ethylene precursor ACC elevated rooting by about 20%. The total content of soluble sugars was proved to be biomarker of rhizogenesis in the studied species. Their concentration was positively correlated with rooting efficiency, while a level of phenolic was positively correlated with the length of regenerated roots, and their number per rosette. The cultivation of the acclimatized material was successfully carried out and was evaluated over three subsequent years. In the third year of cultivation, the plants entered the stage of generative development and most of them bloomed profusely.

Effect of pH, Sucrose Concentrations and Medium States on in vitro Root-ing of Pineapple (Ananas comosus (L) Merr) cv Queen

The effect of medium states (liquid, semi solid, solid), pH (5.0, 5.5, 6.0, 6.5) and sucrose concentrations (10, 20, 30, 40 g/l) on in vitro rooting of pineapple cultured in full strength MS enriched with IBA at 0.5 mg/l were investigated. According to average overall sucrose concentrations, overall pH adjustments, and at each combination of equal sucrose and pH, liquid medium was always super than solid and semisolid. The tallest plantlets (66 to 71 mm) obtained in liquid medium enriched with sucrose at 10 and 20 g/l both adjusted to pH 6.0; sucrose at 20 g/l and adjusted to pH 6.5 and sucrose at 30 g/l and adjusted to pH 5.0. All of the above combinations except sucrose at 20 and pH 6.0 resulted in 100% rooting. Sucrose at 30 g/l and pH 5.0 resulted in two times more (11 roots per shoot) and three times longer roots (39 mm) than the other treatments (5 roots each about 14 mm long). Each rooting parameter had different optimum combinations of medium state, sucrose and pH adjustment. For any parameter, proper pH adjustment could reduce the optimal sucrose enrichment from 30 to 20 and even to 10 g/l. Hence, pH adjustment is suggested as an important approach for reduction of in vitro rooting medium cost.

Strategic Mulching of Trees in Forested Urban Parkland for Rooting Medium Amendment

Trees planted along city streets and in urban parks are subject to many adversities that affect growth and can often result in mortality. The application of organic mulch to the rooting medium of newly planted urban trees has the potential to improve the soil chemical and physical properties necessary for tree root health. This study examined the difference in soil nutrient supply rates (μg/10cm2/28 days) between three areal treatments of wood chips (0.75 m, 1.0m and 1.5 m radii) and before mulch application versus after mulch application using Analysis of Covariance. PRSTM-Probes were inserted into the soil over six 28-day periods to measure the supply rate of bioavailable nutrients (NO3-, NH4+, P, K, S, Ca, Mg, and Cu). Meteorological data and other soil chemical and physical factors were measured and included as covariates in the statistical model. Results indicate that mulching had a significant effect (p<0.05) on P supply rates; supply rates were lower in the reference plots compared to the treatment plots post-mulching. S, Ca, Mg, and Cu supply rates decreased after mulch application; however, the decrease was observed in all plots, which is likely due to temporal variations in plant demand rather than mulching. The wood chips also had a significant impact on buffering fluctuating soil temperatures and reducing soil moisture loss compared to non-mulched plots. The knowledge obtained from this research can be used to improve urban forest management strategies by providing a more in-depth understanding of the prescriptive use of organic mulch.

Strategic Mulching of Trees in Forested Urban Parkland for Rooting Medium Amendment

Trees planted along city streets and in urban parks are subject to many adversities that affect growth and can often result in mortality. The application of organic mulch to the rooting medium of newly planted urban trees has the potential to improve the soil chemical and physical properties necessary for tree root health. This study examined the difference in soil nutrient supply rates (μg/10cm2/28 days) between three areal treatments of wood chips (0.75 m, 1.0m and 1.5 m radii) and before mulch application versus after mulch application using Analysis of Covariance. PRSTM-Probes were inserted into the soil over six 28-day periods to measure the supply rate of bioavailable nutrients (NO3-, NH4+, P, K, S, Ca, Mg, and Cu). Meteorological data and other soil chemical and physical factors were measured and included as covariates in the statistical model. Results indicate that mulching had a significant effect (p<0.05) on P supply rates; supply rates were lower in the reference plots compared to the treatment plots post-mulching. S, Ca, Mg, and Cu supply rates decreased after mulch application; however, the decrease was observed in all plots, which is likely due to temporal variations in plant demand rather than mulching. The wood chips also had a significant impact on buffering fluctuating soil temperatures and reducing soil moisture loss compared to non-mulched plots. The knowledge obtained from this research can be used to improve urban forest management strategies by providing a more in-depth understanding of the prescriptive use of organic mulch.