Browsing by Author "Mokotedi, Mompe Edward Oscar."

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The development of in vitro rooting systems for cold-tolerant Eucalyptus grandis x nitens clones and the assessment of the hydraulic efficiency of roots produced by in vitro vs. cutting propagation.
(1999) Mokotedi, Mompe Edward Oscar.; Watt, Maria Paula Mousaco Deoliveira.; Pammenter, Norman William.
Hybrid clones of the fast-growing Eucalyptus grandis and cold-tolerant E. nitens (GN clones) have been identified by the South African Forestry Industry as being highly suitable for plantations in cold-dry marginal areas. However, one of the main problems regarding their propagation is the difficulty in rooting of cuttings, both in vitro and ex vitro. The aims of this investigation, therefore, were (1) to develop widely applicable and efficient in vitro rooting system(s) for these commercially important clones, and (2) to assess some physiological characteristics of the roots produced. Adventitious shoots (15-20 mm in length) were obtained (l0 shoots/explant) from axillary buds on Murashige and Skoog's (MS) medium containing 0.01 mg.l-1 NAA, 0.01 mg.l-1 IBA and 0.2 g.l-1 FAP. The effect of various medium components, as well as modification of culture environment on in vitro rooting, were investigated. The highest rooting frequencies in clones GN121 (75%) and GN107 (65%) were achieved on l/4 MS with additional 0.22 g.l-1 CaCl2..2H2O and 0.18 g.1-1 MgS04.7H2O, 0.1 mg.l-1 IBA, 0.1 mg.l-1 biotin, 0.1 mg.l-1 calcium pantothenate, 15 g.1-1 sucrose and 4 g.l-1 Gelrite. Best culture conditions were an initial 72-hours dark incubation followed by a 16-hours day/8-hours night photoperiod at a PPFD of 37 µmol.m-2.s-1 and 23°C day/21°C night for seven days, after which the PPFD was increased to 66 µmol.m-2.s-1 at 27°C day/21°C night for 18 days. Towards the development of a more widely applicable in vitro rooting protocol for GN clones, the use of Agrobacterium rhizogenes strains was investigated. Production of transgenic roots was observed on carrot discs and shoots from seedlings of Eucalyptus grandis and E. nitens, but not on shoots of GN clones. Therefore, a method needs to be established for the successful transfer and integration of the Ri plasmid of Agrobacterium into the hybrid plant genome for induction of transgenic roots. The quality of roots produced in vitro and from cuttings was assessed by examination of root anatomy and hydraulic characteristics. Adventitious roots were prepared for measurement of hydraulic conductivity by detopping explants, then filtered, acidified distilled water was drawn through undisturbed potted root systems under partial vacuum, causing no damage to the roots. Initial studies showed that tissue culture-derived roots exhibited a higher specific root mass hydraulic conductivity than those derived from cuttings (6.46 x 10-6 vs. 3.06 X 10-6 g.kPa-1.s-1.g-1 dry root), probably due to root architecture. Curves relating vulnerability to water potential were constructed and both types of roots showed vulnerability to cavitation at high water potentials. Differences were also observed in staining reactions (safranin and fastgreen) which might suggest differences in presence and level of secondary metabolites in these roots at the juvenile stage. Applications of the developed protocols and future research strategies are discussed.
Influence of stockplant management on yield and subsequent rooting of cuttings of cold-tolerant Eucalyptus grandis x E. nitens clones.
(2012) Ziganira, Matabaro.; Bertling, Isa.; Blakeway, Felicity Clare.; Mokotedi, Mompe Edward Oscar.
Clones of the Eucalyptus grandis x Eucayptus nitens (GN) hybrids were produced and selected through the CSIR‟s breeding programmes for colder plantation sites in South Africa. Some GN clones consistently exhibit high and superior pulp properties, which makes them valuable for commercial plantations in South Africa. In nurseries, stockplants are usually seven cm in length and maintained at high (100 x1.5 m-2) planting density. However, rooting frequency varies with season and little is known about the impact of position of cuttings on overall rooting frequency of a clone. The aim of this study was to investigate the effect of size and planting density of stockplants in mini-hedges, on the yield and subsequent rooting of cuttings from various positions of GN clones of known rooting potential (i.e. GN 018B: difficult-to-root and PP 2107: easy-to-root clones). Stockplants (10 cm vs. 20 cm) were established at high (100 x 1.5 m-2) and at low (25 x 1.5 m-2) densities for GN 018B and PP 2107 under commercial nursery conditions in a polyethylene tunnel. Cuttings were harvested every two to three weeks in September-October 2010 (spring), December 2010-January 2011 (summer), April-May 2011 (autumn) and June-July 2011 (winter). The harvested material was 5 – 7 cm in length and the light intensity received by individual stockplants at the two planting density levels was recorded. Harvested cuttings from the three positions (apical, middle and basal shoots) were used for: (i) rooting experiments under nursery conditions, (ii) bio-stimulant analysis using the mung bean bioassay, and (iii) analysis of soluble sugars. Between spring and summer 2010, the two GN clones established at low density yielded a similar number of cuttings, but differences in the rooting frequencies were significant in favour of PP 2107 clone. Similar observations were made at high density in terms of production of cuttings, but the significant differences in the rooting observations were reversed between the clones. The GN 018B clone had low rooting rates in summer under nursery conditions but its tissue extracts promoted higher rooting in the bioassay during that time, when compared to spring. Spring and summer had similar effects on rooting responses of PP 2107 cuttings in nursery and bioassay experiments. For both clones, short stockplants produced fewer cuttings but had a higher rooting frequency than cuttings from tall stockplants, with a high rooting frequency recorded from basal cuttings. Similar results were observed in the bioassay experiments which showed high rooting potential of mung bean hypocotyls cuttings using tissue extracts of PP 2107 cuttings maintained at high planting density. Although apical cutting tissues had high concentration of sugars (i.e. sucrose, glucose and fructose), their rooting rates were usually lower at high and low planting density compared to middle and basal cuttings. Sucrose concentration was the highest sugar present in stockplants grown under low planting density. A higher and lower rooting frequency was also observed in autumn although the two clones responded differently to Quambalaria eucalypti (Sporothrix eucalypti) disease infestations. Position, size and genotype had a significant impact on type and concentration of sugar (i.e. sucrose, glucose and fructose), particularly in PP 2107 clone, although rooting rates in the bioassay did not correlate with sugar contents of Eucalyptus cuttings. High carbohydrate (i.e. soluble sugar) content and auxin concentration increased production and subsequent rooting of cuttings across both clones, particularly in spring. Furthermore, rooting was enhanced by relatively higher light intensity intercepted by individual stockplants and in particular the GN 018B clone. Light intensity in the high and low planting densities caused variation in the rooting frequencies thereby increasing or decreasing soluble sugar and auxin concentrations of the two clones. Light intensity and fertiliser concentration received by tall and short stockplants impacted on endogenous hormone levels thereby increasing or decreasing rooting. High sugar concentration levels of PP 2107 clone increased its susceptibility to fungal infection thereby decreasing its rooting frequency in autumn, as its rooting rates increased in winter. Overall results of the investigation revealed that PP 2107 clone has higher rooting potential than GN 018B clone, in particular at high planting density and if stockplants are not infected by fungal diseases. Higher sugar levels were recorded in spring for PP 2107, although rooting rates of mung bean hypocotyl cuttings were higher in summer for GN 018B, suggesting that sugars have nothing to do with rooting of GN cuttings. Season, planting density and size of stockplants affect the rooting frequency of GN clone. Thus, short stockplants maintained at low and high planting densities are recommended for GN 018B and PP 2107 respectively, although the impacts of fertilisers and pathogen resistance on rooting rates still need to be investigated under similar conditions.
The performance and rooting of eucalyptus grandis x nitens cuttings.
(2007) Murugan, Nelisha.; Watt, Maria Paula Mousaco Deoliveira.; Mycock, David John.; Mokotedi, Mompe Edward Oscar.
Hybrid clones of Eucalyptus grandis and E. nitens (GN) have consistently been shown to be suitable for planting in cold, dry, marginal plantation sites, where they exhibit high yields and superior pulp properties. However, their clonal propagation is hindered by the very poor rooting success of cuttings. The present study aimed at assessing the effect of cutting type, time of year of setting cuttings and Seradix 2 application on rooting and development of cuttings of a commercially important Eucalyptus grandis x Eucalyptus nitens clone (GN107). Cuttings were prepared from clonal hedge coppice at the Mondi Business Paper, Trahar Technology Centre, Hilton. Three cutting types were used (cut at different distances from the node) for each terminal (situated below the apical bud) and non-terminal cuttings. The leaves were trimmed and, for half the cuttings, the base of the stem of cuttings were dipped in Seradix 2 rooting powder (3 g kg-1 4-(indole-3-yl)-butyric acid (IBA). They were then placed into rooting trays (128 inserts/ tray arranged as 8 rows x 16 columns). Seradix 2-treated and Seradix 2-untreated terminal and non-terminal cuttings, cut at, above and below the node (twelve treatments in total) were set in trays with one treatment per column of eight replicates, per tray. There were nineteen trays overall. The trays were filled with peat, perlite and vermiculite (3:3:1) and were maintained in a Mondi greenhouse, with air temperature at 25°C to 27°C (thermostatically activated fans), root zone temperature at 28°C (bed heaters) and 20 second misting at 10 minute intervals (automatic misters). The study was carried out in November 2005, April 2006 and June 2006. In the first experiment, both terminal and non-terminal cuttings were used; thereafter only non-terminal cuttings were used. The plantlet yield was very low, regardless of cutting type, Seradix 2 treatment and the time of year the cuttings were set. The highest plantlet production (12.5%) and rooting frequencies (13.8%) were achieved with non-terminal cuttings treated with Seradix 2. Although not statistically significant, Seradix 2 inhibited shoot production (31.4% for Seradix 2-untreated and 24.2% for treated cuttings). The position at which inserts were cut in relation to the node did not significantly affect the number of plantlets produced and non-terminal cuttings appeared hardier and performed better than terminal cuttings. The time of year of setting cuttings did not have any significant effect on plantlet yield, nonetheless, plantlet yield was highest in cuttings set in November (9.2%) and lowest in April (0.4%). In addition, cuttings set in November (spring), had superior shoot development in terms of the number of cuttings that produced shoots (regardless of root production), shoot length and the mass of shoots relative to root mass. The highest percentages of cuttings that produced roots (regardless of shoot growth) (10%) and the highest number of roots per cutting (2) were part of the June trial. Therefore, cuttings set in June (winter) had superior root development as compared with cuttings set in November (spring) or April (autumn). In all of the studies, three rooting patterns were observed in cuttings: roots produced only from the cut area only (type 1), only from the sides of the stem (type 2) and from both sites (type 3). Non-terminal cuttings treated with Seradix 2 showed a higher incidence of types 2 and 3 rooting patterns than the terminal cuttings. Seradix 2 application increased the prevalence of types 2 and 3 rooting patterns. Although not statistically different, cuttings dipped 2.5 cm into Seradix 2 produced more types 2 and 3 rooting patterns than cuttings dipped at the abaxial end only. Light microscopy of stem sections of cuttings indicated that roots appeared to originate from the xylem archs as well as from the cambium. The collected data indicate that it is necessary to continue research towards improving the efficiency of plantlet production of GN107 via cuttings. It appears that cuttings of this clone may be set throughout the year and that terminal cuttings should be avoided. In addition, the present practice at the Mondi Hilton nursery of treating cuttings with Seradix 2 needs to be reconsidered as although it increases rooting, it does not increase plantlet production due to its apparent inhibitory effect on shoot development.
Survival and rooting of selected vegetatively propagated Eucalyptus clones in relation to supplied auxin.
(2013) Rambaran, Natasha.; Watt, Maria Paula Mousaco Deoliveira.; Mokotedi, Mompe Edward Oscar.; Nakhooda, Muhammad.
Eucalyptus spp. and hybrids dominate the global plantation forestry industry, and vegetative propagation through cuttings is the preferred method for their commercial use. However, the cuttings of some species and hybrids show recalcitrance to rooting. The first aim of this study was to improve percentage rooting of three clones of E. grandis x E. nitens (Clones 1, 2 and 3) identified by a commercial nursery as having variable rooting abilities. The second was to relate their rooting responses as cuttings to their rooting responses in vitro. Minicuttings (3.5 – 4 cm in length) (hereafter referred to as cuttings) were subjected to commercial nursery propagation practices. Initial results revealed that in the absence of exogenous plant growth regulators (PGRs), soft (juvenile, thin diameter) cuttings survived (87 – 95%) and rooted (29 – 32%) significantly better than hard (mature, thick diameter) ones (62 – 71% survival and 2 – 8% rooting). This validated the use of soft cuttings by the nursery and all subsequent studies were conducted with soft cuttings. The other nursery practice of applying the commercial rooting powder Seradix 2 (3 g kgˉ¹ indole-3-butyric acid [IBA]) adversely affected the survival and subsequent rooting of cuttings of Clones 1 and 2. Ensuing studies investigated: 1) the effect of mode of IBA application (powder vs. liquid); 2) concentrations of Seradix (0, 0.5, 1, 2 and 3 g kgˉ¹ IBA), applied at initial placement of cuttings and two weeks later; and 3) the influence of season on the survival and subsequent rooting of cuttings. Results showed that regardless of the mode of application, IBA significantly reduced percentage survival and rooting in cuttings of Clones 1 and 2. The delayed application of Seradix, two weeks after cuttings were initially set, resulted in a higher percentage survival and rooting than when cuttings were supplied with Seradix at initial placement. Nevertheless, the best survival for Clones 1, 2 and 3 (95%, 99% and 71%, respectively) and rooting (83%, 64% and 47%, respectively) occurred in the absence of Seradix. In addition, the survival and rooting of cuttings were seasonally variable, with particularly low rooting during winter (e.g. for Clone 1, 32%) when compared with summer (e.g. for Clone 1, 83%). Shoots from all the clones were multiplied in vitro, followed by elongation on either of two media (E1= kinetin, α-naphthalene acetic acid [NAA] and IBA; E2 = kinetin and indole-3-acetic acid [IAA]), and then rooting on 0, 0.1 or 1.0 mg 1ˉ¹ IBA. The latter were selected to typify the range of Seradix concentrations used for the cuttings (i.e. no IBA, low and high IBA concentrations). For all three clones, shoots elongated on E1 or E2 displayed high survival (> 80%) but failed to root without IBA in the rooting medium. For Clones 1, 2 and 3 the best in vitro survival (80%, 100% and 100%, respectively) and rooting (40%, 75% and 40%, respectively) occurred when shoots were elongated on E2 and rooted on 0.1 mg 1ˉ¹ IBA. However, 1.0 mg 1ˉ¹ IBA in the rooting medium severely inhibited survival (0 – 50%), irrespective of the clone or the elongation treatment used. Overall, cuttings demonstrated the best survival and rooting in the absence of exogenous IBA, which suggested that sufficient endogenous auxin was present within the shoots for successful root induction. The application of exogenous IBA may have disrupted the cuttings’ endogenous PGR balance resulting in an inhibition of survival and rooting. In vitro shoots required a low concentration of IBA (0.1 mg 1ˉ¹) in order to counteract the antagonistic effect of cytokinins that were supplied during the multiplication and elongation culture stages, and promote rhizogenesis. Essentially, both cuttings and in vitro shoots demonstrated adverse survival and rooting responses when subjected to excessively high IBA concentrations.