This study focused on induction of somatic embryogenesis of Magnolia spp. and cultivars utilizing leaf and seed (immature and mature) tissues with attempted micropropagation experiments. In a preliminary experiment, direct embryo regeneration was successful in a single leaf tissue of M. ‘Yellow Bird’. After various micropropagation experiments, microshoot proliferation rates decreased. As a result of minimal leaf material, mature seeds were utilized but had contamination issues. Subsequent experiments utilized immature seeds. M. ‘Leonard Messel’ and M. stellata had significantly greater embryo regeneration rates and M. ‘Rosea’, M. stellata, and M. kobus had greater callus induction rates. Woody Plant Medium had significantly greater rates of embryo regeneration as compared to Yellow Poplar medium. Further experimental measures including various collection times of immature seeds are necessary for an efficient regeneration protocol to support potential research utilizing floral-inducing genes to induce rapid breeding cycles for selection of magnolias with diverse floral characteristics.

No description available.

No description available.

Dormant bud and actively growing shoot tip explants of B. uber were cultured on Murashige & Skoog (MS) and modified woody plant (WPM) basal salts supplemented with organic nutrients and benzylaminopurine (BAP). Shoot tip explants were morphogenically more responsive than dormant bud explants. The modified WPM proved to be better than MS medium for the establishment and proliferation of B. uber cultures. Shoot proliferation was greatest at 8.9 μM BAP. However, BAP at 4.4 μM produced the greatest number of shoots > 10 mm. Indolebutyric acid (IBA) was more effective than naphthaleneacetic acid (NAA) for rooting of microcuttings in vitro. Rooting was best with a low level of IBA (2.5 μM) in the culture medium. Ex vitro rooting of microcuttings using a conventional potting mix of sand-peat-perlite (1:2:1 v/v/v) was superior to all in vitro rooting treatments. Plantlets were acclimated successfully to greenhouse conditions. Genotype differences with regard to both rooting and shooting abilities were observed. / Master of Science

Thesis (M. Sc. (Agricultural Science)) -- University of Limpopo, 2001 / Refer to document

Date palms (Phoenix dactylifera L.) and papayas (carica papaya L.) are two commercially important plantation crops. Their economic potential in South Africa and worldwide is increasing. However, due to disease, pests and socio-economic reasons, planting material is in short supply. Micropropagation provides a method for rapidly propagating selected superior cultivars for commercial and environmental interests. A satisfactory process for the regeneration of elite cultivars should result in individuals phenotypically and genetically identical to the explant from which they were derived. However, due to somaclonal variation generated during in vitro culture, the true-to-typeness is questionable. For this reason a southern African survey for off-types on date palms produced using somatic embryogenesis was conducted. Plant growth variations such as leaf variegation, seedless fruit, broad leaves, compact growth habit and parthenocarpic fruit were recorded and possible explanations for each phenomenon given. Factors influencing the date palm initiation process such as decontaminating agents, plant growth regulators, explant type and nurse cultures were investigated. A double decontamination process with 2.6% and 1.3% sodium hypochlorite was most effective at reducing contamination. Alternative plant growth regulators, TIBA and NAA were ineffective as a substitute to 2,4-D for somatic embryogenesis. The size of the explant and "nurse cultures" played an important role in explant growth and initiating callogenesis. A "nurse culture" reduced the time in culture significantly. The problem areas in the three commercial tissue culture techniques used for date palms were outlined. In the second part of the study, factors influencing initiation, multiplication and rooting of papaya were determined. Presoaking with antibiotic, Rifampicin, and various fungicides had a positive effect on decontaminating papaya explants, while Bronocide™ had little effect. Various methods and materials were used to optimize papaya multiplication and rooting in vitro. The growth and multiplication of papaya was optimal at 50 g l ¯¹ sucrose. Gelling agent, Gelrite, increased multiplication rates significantly but had a negative effect on overall growth causing plants to become vitrified. The addition of activated charcoal reduced vitrification but also reduced multiplication rate. Activated charcoal greatly improved overall growth of papaya and reduced leaf senescence. No vitrification was observed in multiplying papaya cultures where agar and Gelrite combinations were used, but multiplication rate was reduced compared to cultures grown on Gelrite alone. callus removal from the bases of papaya at subculturing reduced multiplication rate and influenced elongation, growth and leaf senescence. Lower concentrations magar and Gelrite improved rooting percentages, but did not provide good support. Damaged roots and lower rooting percentages were observed on plantlets treated with IBA for four weeks compared to those exposed for only two days. A one hour pulse with a higher concentration (5 mg l ¯¹) of IBA greatly improved rooting percentage and further eliminated a second subculture onto an IBA-free medium after two days. Good, strong roots with root hairs were produced on vermiculite medium containing equal volumes of DS salts and vitamins. Modified lids with cotton-wool plugs also reduced leaf abscission. In vitro grafting using stericrepe proved impractical, while grafting in vitro unrooted papaya plants onto ex vitro seedlings was more successful, using wedge and slant grafts. Grafts sealed with pegs and Parafilm™ were less effective. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 2000.

Banana production ranks fifth behind cereals as a food crop and has potential, along with other major crops, to feed the world's increasing population. Globally, continuous efforts and techniques including the use of plant tissue culture (PTC) have been devised for increasing the production of several Musa species. The choice of cytokinin (CK) is one of the most critical factors in developing a successful PTC protocol. Since the discovery of topolins as naturally occurring aromatic CKs, they have emerged as genuine alternatives to the long serving CKs (benzyladenine = BA, zeatin = Z and kinetin = KIN) in PTC. Globally, the past 15 years has witnessed a surge in the use of topolins and their derivatives in research laboratories. Topolins have demonstrated great potential during culture initiation and protocol optimization as well as for counteracting various in vitro induced physiological disorders in some species. In terms of general physiology (growth, phytochemical and photosynthetic pigment contents as well as genetic fidelity), the topolins were compared with BA using 'Williams' bananas with minimal residual exogenous CK carry-over effects. The five topolins tested were meta-Topolin (mT); meta-Topolin riboside (mTR); meta-Methoxy topolin (MemT); meta-Methoxy topolin riboside (MemTR) and meta-Methoxy topolin 9-tetrahydropyran-2-yl (MemTTHP). Based on evidence of potential CK- and auxin-like activity of smoke-water (SW) and karrikinolide (KAR1) at low concentrations, a similar comparative study involving both compounds and mT was performed. For a further understanding of banana physiology in vitro, the effect of supplementing either mT- or BA-requiring cultures with roscovitine (a cyclin-dependent kinase and N-glucosylation inhibitor) and INCYDE (an inhibitor of CK degradation) on the endogenous CK profiles was investigated. In addition, greenhouse experiments geared towards improving the acclimatization competence of tissue-cultured banana plantlets via application of different concentrations of SW and vermicompost leachate was conducted. Sterile shoot-tip explants were cultured on modified Murashige and Skoog (MS) media supplemented with 10, 20 or 30 μM of the tested CKs for 42 days while rooting experiments involved the use of classic auxins as well as SW and KAR1. Apart from 10 μM BA and 30 μM MemTTHP treatments, the number of shoots produced with all the CK treatments were significantly higher than the control. Treatment with 30 μM mT resulted in the highest number of shoots (7.3±1.0) which is an indication of the requirement of exogenous CK for increased shoot proliferation in 'Williams' bananas The use of 10 μM MemTTHP had the least root inhibitory effect during the shoot proliferation phase. As an indication of the toxicity of applied CK, MemT- and MemTR-regenerants were the most deformed while mTR-regenerated plantlets demonstrated the best quality across all the CKs tested. In mT- and BA-derived shoots, SW and KAR1 significantly increased the number and length of roots compared to the control. During the rooting phase, topolin treatments produced more off-shoots than BA-treated ones which inevitably improved the overall number of regenerated shoots. Total phenolic levels were highest in 10 μM mT- and 30 μM MemTTHP-treated plantlets detected in the aerial and underground parts, respectively. It is interesting that in the underground parts, 10 μM mT resulted in the production of the highest amount of proanthocyanidins which was approximately five-fold higher than in the control plants. On the other hand, 10 μM MemTTHP-treated plantlets had significantly higher total flavonoids within the aerial parts. In view of the stimulation of secondary metabolites in the majority of the CK-treated plantlets, the current results indicate the role of the type and concentration of applied CK as potential elicitors in PTC. Generally, the maximum photosynthetic pigment content was attained between 40-50 days. The control plantlets had the highest pigment content (1150 μg/g FW) while 10 μM MemTTHP had the best pigment stimulatory effect among the tested CKs. Nevertheless, in vitro propagation of banana devoid of CKs is not a practical option due to low shoot proliferation rates. Scanning electron microscopy (SEM) of the foliar surface showed that the stomatal density was highest in 10 μM MemTTHP-treated and lowest in 10 μM MemTR-treated plantlets. Prolonging the culture duration as well as increasing CK concentrations reduced the pigment content. However, the drastic breakdown in chlorophyll pigments beyond 50 days was slightly inhibited by the presence of mT, mTR, MemTTHP and BA compared to the control. Current findings indicate the potential anti-senescence activity of the topolins such as mT, mTR and MemTTHP under in vitro conditions. This study articulates that the right choice and concentration of CKs applied during in vitro propagation may alleviate photomixotrophic-induced physiological stress that usually accompanies the transfer of plantlets to ex vitro conditions. Findings indicate that the effect of subculturing contributed significantly to the higher rate of variation in 'Williams' bananas in vitro. The presence of CK in the culture media apparently aggravated the stress on the explants as indicated in the relatively higher percentage polymorphic bands compared to the controls. Among the tested CKs, the use of mTR and MemTTHP caused the least detrimental effect on the regenerants while mT-treated plantlets had the most polymorphic bands. Hence, it is recommended that subculturing cycles from the initial explant establishment should be limited to a maximum of five. The use of SW and KAR1 improved the level of photosynthetic pigment and phenolic compounds in the micropropagated bananas. However, they had a negative effect on shoot proliferation; hence their inclusion is more desired when used at the rooting phase of micropropagation. Perhaps, these compounds could be used in conjunction with auxin to increase the number of roots prior to the acclimatization stage. The enhanced photosynthetic pigment level resulting from addition of SW and KAR1 would also play a vital role during acclimatization of the micropropagated plants. The present finding serves as an alternative approach, available to researchers for improving the quantity of secondary metabolites in micropropagated plants. The highest regeneration rate (93%) was observed in BA + roscovitine treatment while mT + INCYDE-treated plantlets produced most shoots. Treatment with BA + roscovitine had the highest shoot length and biomass. Although not significant, there was more proanthocyanidins in BA + roscovitine treatments compared to the treatment with BA alone. On the contrary, total phenolics were significantly higher in mT + roscovitine treatment than in the mT-treated regenerants. The presence of roscovitine and/or INCYDE had no significant effect on the photosynthetic pigments of the banana plantlets. Forty-seven aromatic and isoprenoid CKs categorized into nine CK-types were detected at varying concentrations. The presence of mT + roscovitine and/or INCYDE increased the levels of O-glucosides, while 9-glucosides remained the major derivative in the presence of BA. Generally, the underground parts had higher CK levels than the aerial parts; however the presence of INCYDE increased the level of CK quantified in the aerial parts of both CK treated plantlets. Apparently, the presence of INCYDE serves to enhance transportation of the CK towards the aerial regions. From a practical perspective, the use of roscovitine and INCYDE in PTC could be crucial in the alleviation of commonly observed in vitro-induced physiological abnormalities. Soil drenching with SW significantly increased the root length (1:1000 and 1:500 dilutions) as well as fresh and dry weight (1:1000; 1:500 and 1:250 dilutions) when compared to foliar application. Vermicompost leachate (1:10 and 1:5 dilutions) significantly enhanced the shoot length, root length, leaf area and dry weights. Vermicompost leachate (1:20; 1:10 and 1:5 dilutions) also significantly increased the number of off-shoots. The positive effect on rooting is beneficial for acclimatization and establishment of tissue-cultured banana plantlets in nurseries and subsequent transfer to the field. However, field trials will be necessary to substantiate the effects demonstrated by these compounds. In an attempt to contribute to improving banana micropropagation, the current findings provide additional evidence on the increasing advantage of topolins over BA. Nevertheless, some detrimental physiological effects observed with some of the topolins (for example, MemT and MemTR) are clear indication that they should not be taken as a panacea in PTC. Besides optimizing efficient PTC protocols through stringent choice of CKs, other associated physiological and metabolic events taking place in culture during the optimization process need more in-depth investigation. In addition to contributing towards the better understanding of the mode of action of these CKs, such an approach will help solve associated physiological and developmental problems in vitro. / Thesis (Ph.D.)-University of KwaZulu-Natal, Pietermaritzburg, 2012.

The effect of activated charcoal, autoclaving and culture media on sucrose hydrolysis in tissue culture media was investigated. Activated charcoal acidified an aqueous sucrose (5%) solution and culture media by about 1 to 2 units after autoclaving . Sucrose hydrolysis in tissue culture media and/or aqueous sucrose (5%) solutions containing activated charcoal (buffered to pH 5.8) was dependent on both the hydrogen ion concentration (pH) and autoclaving. After autoclaving, 70%, 56% and 53% sucrose hydrolysis were respectively recorded in a 5.0% sucrose solution, Murashige and Skoog (MS) and Gamborg B5 (B5) liquid media in the presence of 1.0% activated charcoal, added before autoclaving . In the absence of activated charcoal, autoclaving resulted in about 20% of the sucrose being hydrolysed The adsorption of 2, 4-dichlorophenoxyacetic acid (2,4-D) by activated charcoal from methanol and aqueous solutions was determinated using HPLC. The amount of the added 2,4-D decreased in both methanol and aqueous solutions in the presence of activated charcoal, compared with those in the absence of activated charcoal. In methanol and aqueous solutions, activated charcoal used at the level of 0.1% significantly reduced 2,4-D. About 68.4% and 60.9% respectively of the added 2,4-D was adsorbed by activated charcoal (1.0%) from these solutions. The changes of inorganic elements in MS-salt solutions, in the presence of activated charcoal, were analysed by SEM-EDX. The concentrations of magnesium (Mg), calcium (Ca), iron (Fe) and zinc (Zn) deceased in the presence of activated charcoal, while the concentrations of potassium (K), copper (Cu), manganese (Mn), phosphorus (P), and sulphur (S) increased in the MS salt solution in the presence of activated charcoal compared with no activated charcoal in the medium. This suggests that activated charcoal adsorbed calcium, magnesium, iron and zinc and released copper, manganese, phosphorus and sulphur. Rooting occurred when 7-day-old seedling hypocotyls of Daucus carota L. Cape Market were placed on MS medium supplemented with 2,4-D, and IAN/NAA in the presence of activated charcoal. Hypocotyls did not produce roots on the 2,4-D containing media in the absence of activated charcoal. The roots were produced polarly on the NAA/IAA-containing media in the presence of activated charcoal. No-polarity of root formation was observed on media supplemented with NAA/IAA without activated charcoal. Different responses of hypocotyls to a series of 2,4-D concentrations (0.5, 1.0, 3.05.0, 8.0, and 10.0 mg l ¯¹) were observed on media supplemented with 0.02, 0.1 and 0.5% activated charcoal. In the NAA/IAA containing media in the presence of activated charcoal, root number per hypocotyl decreased. Root number perhypocotyl, on the media supplemented with NAA and IAA, increased when hypocotyls were pre-cultured on MS medium supplemented with 2,4-D (1.0 mg l ¯¹) for 2-3 days. When hypocotyls were pre-cultured on a 2,4-D containing MS medium for 5 days, embryos emerged from the hypocotyls directly on the medium supplemented with 2,4-D in the presence of activated charcoal. Addition of activated charcoal to MS medium supplemented with 2,4-D resulted in somatic embryogenesis of Daucus carota. Somatic embryos were not formed on the medium in the absence of activated charcoal. In suspension culture, the incorporation of 0.01 to 1.0% concentrations of activated charcoal to the MS medium, irrespective of 2,4-D, increased the number of somatic embryos produced. The maximum number of somatic embryos were produced with 1.0% activated charcoal. Further development of embryos of Daucus carota occurred on the media in the presence of activated charcoal, and the embryos subsequently regenerated normal plantlets. Abnormal somatic embryos followed the addition of 3.0% activated charcoal to the medium. / Thesis (Ph.D.)-University of Natal, Pietermaritzburg, 2000.

Field and greenhouse studies were conducted in 1986 and 1987 to determine the impact of disturbance of microphytic soil crust on emergence of two warm season grass species. In the greenhouse, emergence of seedlings sown on the top of undisturbed crust, under the crust, on disturbed crust, and on bare soil was studied. The mean number of seedlings of "Cochise" lovegrass (Eragrostis lehmanniana x Eragrostis tricophora) and Kleingrass (Panicum coloratum) was higher on disturbed field plots than undisturbed plots. Disturbance reduced the microphytic crust cover, and the crust did not recover throughout the growing season. Presence of seedlings in samples of litter washed off the plots and observation on ants showed that all the seeds sown did not remain on the plots during the growing season. In greenhouse flats the emergence of seedlings of both species was greater on disturbed microphytic crust and bare soil than when seeded on top of or under an intact crust. Undisturbed crust restricted penetration of roots and shoots when seeds were placed on or under the crust.

Thesis (MSc (Botany and Zoology))--Stellenbosch University, 2011. / ENGLISH ABSTRACT: See full text for abstract / AFRIKAANSE OPSOMMING: sien volteks vir opsomming