Browsing by Author "Mulaudzi-Masuku, Takalani"
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Item AtNOGC1 protein bioelectrode for the determination of stress signalling molecules - Nitric Oxide (NO), Carbon Monoxide (CO) and Calcium ion (Ca2+)(University of the Western Cape, 2018) Tshivhidzo, Tsumbedzo Tertius; Mulaudzi-Masuku, Takalani; Iwuoha, Emmanuel; Ajayi, FanelwaIt has been estimated that the world population will reach about 10 billion by the year 2050 and in order to accommodate the increased demand of food, the world agricultural production needs to rise by 70 % in the year 2030. However, the realisation of the goal in food production is hindered by limited arable land caused by urbanisation, salinisation, desertification and environmental degradation. Furthermore, abiotic and biotic stresses affect plant growth and development, which lead to major crop losses. The long term goal of this study is to improve food security by producing genetically engineered agricultural crops that will be tolerant to diverse stresses. This research aims at developing stress tolerant crops through the determination of important signalling molecules and second messengers, such as nitric oxide (NO), carbon monoxide (CO) and calcium ion (Ca2+), which can bind to plant proteins such as AtNOGC1 in order to induce stress tolerance in plants.Item Characterization and expression analysis of heme oxygenase genes from sorghum bicolor(SAGE Publications, 2019) Mulaudzi-Masuku, Takalani; Ikebudu, Vivian; Muthevhuli, MphoHeme oxygenases (HOs) have a major role in phytochrome chromophore biosynthesis, and chromophores in turn have antioxidant properties. Plant heme oxygenases are divided into the HO1 sub-family comprising HO1, HO3, and HO4, and the HO2 sub-family, which consists of 1 member, HO2. This study identified and characterized 4 heme oxygenase members from Sorghum bicolor. Multiple sequence alignments showed that the heme oxygenase signature motif (QAFICHFYNI/V) is conserved across all SbHO proteins and that they share above 90% sequence identity with other cereals. Quantitative real-time polymerase chain reaction revealed that SbHO genes were expressed in leaves, stems, and roots, but most importantly their transcript level was induced by osmotic stress, indicating that they might play a role in stress responses. These findings will strengthen our understanding of the role of heme oxygenases in plant stress responses and may contribute to the development of stress tolerant crops.Item The effect of chitosan in the germination and growth of sorghum bicolor under salt stress(University of the Western Cape, 2022) Nkuna, Mulisa; Mulaudzi-Masuku, TakalaniThe agricultural sector has been facing enormous challenges including reduced crop yield and increased food production demand to cater for the growing population. Crop production is severely affected by abiotic and biotic factors, however, abiotic stresses especially salinity is a major factor that has contributed to more than 50% loss of important crops including rice, wheat, and maize. The use of fertilisers and pesticides played a vital role in improving crop yield and quality throughout the seasons. However, in the last decade, there has been a rising concern about their negative effects on human health and the environment due to their overuse.Item Exogenous carbon monoxide and jasmonic acid mitigate salt stress in Sorghum bicolor(University of the Western Cape, 2022) Sias, Gershwin; Mulaudzi-Masuku, TakalaniThe agricultural sector plays an important role in the world economy. Expanding research is piloting its improved contribution to the economic sector. Despite the economic gain, the increasing population has already put a strain on agriculture, but the effects of abiotic stresses are severely affecting crop production globally. Abiotic stress is not only affecting the immediate plants, but further inhibits soil fertility, which will affect future crop productivity. The damaging effects of salt stress is only expected to intensify, which further obscures the future prospect of food security. Salt stress management should be prioritised to alleviate food insecurity. Understanding the mechanism involved in conferring salt stress tolerance to plants is fundamental in the establishment of crops with improved tolerance. Sorghum bicolor is one of the most important crops globally that is moderately tolerant to drought and salt stress thus in addition to the availability of its genome, sorghum can be the perfect model crop to further elucidate the mechanism involved in salt stress tolerance.Item Exogenous carbon monoxide and jasmonic acid mitigate salt stress in Sorghum bicolor(University of the Western Cape, 2022) Sias, Gershwin; Mulaudzi-Masuku, TakalaniThe agricultural sector plays an important role in the world economy. Expanding research is piloting its improved contribution to the economic sector. Despite the economic gain, the increasing population has already put a strain on agriculture, but the effects of abiotic stresses are severely affecting crop production globally. Abiotic stress is not only affecting the immediate plants, but further inhibits soil fertility, which will affect future crop productivity. The damaging effects of salt stress is only expected to intensify, which further obscures the future prospect of food security.Item Identification and characterisation of sorghum bicolor heme oxygenase-1 (sbho1) gene and its role in conferring biotic and abiotic stress tolerance to plants(University of the Western Cape, 2019) Ikebudu, Vivian; Mulaudzi-Masuku, TakalaniBiotic and abiotic stresses such as herbivore attack, pathogen infection, salinity, drought, cold, heat and UV radiation, are unfavourable conditions that affect plant growth and productivity globally (Fedoroff et al., 2010). Agricultural productivity and food security are severely limited by salinity, drought and temperature. These environmental factors occur as a result of climate change, which is predicted to cause an increase in the occurrence of severe weather and may stimulate increased damage to plants (Zhu, 2016). Pathogen and herbivore attacks caused a 31 - 42 % loss in crop yield and about 6 - 20 % post-harvest loss (Tesfaw & Feyissa, 2014). Estimates have shown that over 50 % loss of crop yield worldwide is as a result of abiotic stresses (Lobell et al., 2011) and their severity would lead to an increase in loss of arable land yearly. Population growth and migration to these arable lands further increases the issue of land desiccation and by 2050, food production will have to be increased by at least 70 % to meet the growing demand for quality, nutritious and sustainable food. Various biotic and abiotic stresses lead to the increased production of reactive oxidative species (ROS) in plants that cause damage to proteins, lipids and nucleic acids, resulting in cell death (Choudhury et al., 2017).Item Identification and characterization of a heat-inducible Hsp70 gene from Sorghum bicolor which confers tolerance to thermal stress(Springer, 2015) Mulaudzi-Masuku, Takalani; Mutepe, Rendani Daphney; Mukhoro, Ofhani ChristopherThis study describes the first detailed molecular characterization of the heat shock protein 70 (Hsp70) gene from Sorghum bicolor, MN1618 designated as SbHsp70-1. The full-length cDNA of SbHsp70-1 consists of 2524 bp with a 1950 bp open reading frame, which encodes a protein of 649 amino acids. SbHsp70-1 is a cytoplasmic protein with high homology to other plant Hsp70s, especially grain crops. Recombinant SbHsp70-1 was able to bind and hydrolyse ATP in a dose-dependent manner, suggesting that SbHsp70-1 functions as an ATPase.Item Investigation of Guayule's Agronomic Performance and Agro-processing in South Africa(University of the Western Cape, 2017) Mutepe, Rendani Daphney; Ndimba, Bongani; Cornish, Katrina; Mulaudzi-Masuku, TakalaniGuayule (Parthenium argentatum) is grown for its high quality hypoallergenic natural rubber latex production. The plant is native to the Chihuahua desert of North America and successful trials have been done in Europe, the United States of America (USA) and the Republic of South Africa (RSA). It is already undergoing industrial scale development in the USA producing good quality rubber products with impressive stretchiness and strength. The performance of guayule lines AZ1, AZ2, AZ3, AZ4, AZ5, AZ6, OSU1, and 11591 was investigated in the Eastern and Western Cape regions in RSA with an ultimate goal of leading to commercial production. To ensure continuous supply of plants, and avoiding documented seed dormancy issues, a micropropagation protocol was established using the OSU1 guayule line. In addition, laboratory scale latex extraction was accomplished using the Waring blender method with KOH pH 11 buffer and the amount of latex was quantified using the 1 ml latex quantification method. The extracted latex homogenate from the different guayule lines was pooled and purified using Sodium Carboxymethyl Cellulose. The efficiency of different molecular weights (90 000, 250 000 and 700 000) of Sodium Carboxymethyl Cellulose was determined in creaming guayule latex at room temperature and 4 degrees celcius. The optimal creaming results were incorporated into creaming latex extracted from the different guayule lines during this study.Item Investigation of the role of AtNOGC1, a guanylyl cyclase protein in response to abiotic and biotic stress(University of the Western Cape, 2018) Muthevhuli, Mpho; Mulaudzi-Masuku, Takalani; Iwuoha, Emmanuel; Donaldson, LaraAgricultural production is one of the most important sectors which provide food for the growing world population which is estimated to reach 9.7 billion by 2050, thus there is a need to produce more food. Climate change, on the other hand, is negatively affecting major global crops such as maize, sorghum, wheat and barley. Environmental factors such as salinity, drought, high temperatures and pathogens affect plant production by oxidatively damaging the physiological processes in plants, leading to plant death. Poor irrigation used to combat drought result in salinasation, which is estimated to affect 50% of arable land by 2050. Plants have developed several mechanisms that protect them against stress and these include overexpression of stress responsive genes and altered signal transduction to change the expression of stress responsive genes, among others. Cyclic 3’5’ guanosine monophosphate (cGMP), a second messenger that is synthesised by guanylyl cyclase (GC), transmit signals to various cellular functions in plants during plant development, growth and response to abiotic and biotic stresses. Arabidopsis thaliana nitric oxide guanylyl cyclase 1 (AtNOGC1) is a guanylyl cyclase which upon activation by nitric oxide (NO) leads to the production of more cGMP. Cyclic GMP further activates protein kinases, ion gated channels and phosphodiesterase which mediate response to various stresses. In this project the role of AtNOGC1 was investigated in response to abiotic and biotic stresses through analysis of its evolutionary relationships, promoter, gene expression and functional analysis via the viability assays in Escherichia coli (E.coli). Phylogenetic tree, exon-intron structure and conserved motifs were analysed using the Molecular Evolutionary Genetics Analysis (MEGA V.7), Gene Structure Display Server 2.0 (GSDS 2.0), and Multiple Expectation Maximisation for Motif Elicitation (MEME) tools respectively. AtNOGC1’s gene expression was analysed by the Real-Time Quantitative Reverse Transcription Polymerase Reaction (qRT-PCR), whereas functional analysis was carried out using the cell viability (liquid and spot) assays to determine its ability to confer stress tolerance to E. coli.Item Mitigation of salinity stress using exogenously applied molybdenum in sorghum bicolor(University of the Western Cape, 2022) Mabiya, Thembeka Confidence; Mulaudzi-Masuku, TakalaniThe agricultural sector is the main producer of food throughout the world. However, the constant changes in environmental conditions, such as extreme weather, droughts, and salinity have impacted this sector negatively over the years. These stresses cause nutritional imbalance, delayed seed germination and decreased growth resulting in reductions in crop yield and hence affect food prices. The food and agricultural organization (FAO), reported that the average increase rate in crop production is below the amount required to cater for the growing population. Thus, to meet the food demands, discovery of several strategies to improve crop growth and yield under severe environmental conditions are imperative.Item Recombinant expression of cytochrome P450-2D6 and its application in tamoxifen metabolism(University of the Western Cape, 2018) Edwin, Munyai Vukosi; Mulaudzi-Masuku, TakalaniBreast cancer is regarded as the most common form of cancer in women and it comprises of approximately 23 % of female cancers, while affecting women at any age range. For oestrogen receptor positive patients, tamoxifen is used as a prescribed medication for breast cancer therapy. However, tamoxifen in its natural form is not active to achieve the required treatment and prevention of breast cells proliferation. Since tamoxifen is a prodrug, it need to be converted into its active form, endoxifen, for which it is achieved by the action of the cytochrome P450 enzymes. Cytochrome P450 2D6 (CYP2D6) is a member of cytochrome P450 enzymes for which are superfamily of heme enzymes characterised by their ability to catalyse the oxidative reactions of compounds, including the pathway of tamoxifen metabolism. However, due to polymorphism that lead to inactive phenotypes of CYP2D6 in this gene, there is a challenge of diagnosing if a patient can metabolise tamoxifen or not. The current diagnostic tool, Amplichip CYP450, for CYP2D6 is based on genotypes, and it lead to uncertainness as to whether the presence of functionalCYP2D6 alleles of CYP2D6 may lead to coding of active protein, thus leading to wrong treatment measures and overdose of tamoxifen. Electrochemical techniques have provided reliable, simple, quick, and sensitive methods for the determination of drug metabolism by enzymes. Therefore, it is important to develop a CYP2D6 phenotype-based sensor to detect and tell whether a particular individual can metabolise the drug or not.Item Signalling molecule “calcium” improves germination and growth of Sorghum bicolor seedlings under salt stress(University of the Western Cape, 2021) Hendricks, Kaylin; Mulaudzi-Masuku, TakalaniAbiotic stress, mainly in the form of extreme temperatures, drought and salinity has caused major crop losses worldwide, putting a severe strain on agriculture. Salinity severely limits plant growth and productivity and affects all aspects of the plant’s development including the most crucial stage; germination. This study investigated the effect of salt (NaCl) stress on Sorghum bicolor seedlings and the role of exogenously applied calcium (Ca2+) to ameliorate the effects of salt stress during germination. Sorghum seeds were germinated in the presence and absence of various NaCl (100, 200 and 300 mM) and Ca2+ (5, 15 and 35 mM) concentrations. Several assays including physiological (germination and growth assays), biochemical (osmolytes and oxidative stress markers), anatomical (epidermal and xylem layers) and expression profiles of key genes [antioxidant (SbSOD, SbAPX2 and SbCAT3), Salt Overly Sensitive (SbSOS1, 2 and 3) pathway enzymes and the vacuolar Na+/H+ exchanger antiporter2 (SbNHX2)] were investigated. Salt stress delayed germination and negatively affected growth as observed by the reduced root and shoot length and decreased fresh and dry weight. There was an increase in proline content and oxidative stress markers (H2O2 and MDA) under salt stress. Oxidative stress resulted in damage to the epidermal and xylem layers as observed on Scanning Electron Microscopy (SEM) images. Quantitative real-time polymerase chain reaction revealed that salt stress induced the expression of SbAPX2, SbCAT3 and SbSOS1 genes, whereas SbSOD4A, SbSOS2, SbSOS3 and SbNHX2 genes were not affected by salt. Exogenous application of Ca2+ counteracted the harmful effects of salt stress by improving germination efficiency, promoting seedling growth, reducing oxidative damage and the Na+/K+ ratio, indicating the protective effect. Ca2+ also effectively protected the epidermis and xylem layers from the severe damage caused by salt stress. In the presence of Ca2+ the expression of SbAPX2 and SbCAT3 was reduced except for the SbNHX2 gene, which increased by 65-fold compared to the control. The results obtained suggests that sorghum is able to respond to salt stress by inducing osmolytes, the antioxidant defence system as well as the SOS pathway. Furthermore, 5 mM Ca2+ was determined as the optimum Ca2+ concentration required to enhance sorghum’s tolerance to salt stress.Item Spectroelectrochemical graphene-silver/zinc oxide nanoparticulate phenotype biosensors for ethambutol and pyrazinamide(University of the Western Cape, 2019) Tshoko, Siphokazi; Ajayi, Fanelwa; Iwuoha, Emmanuel; Mulaudzi-Masuku, TakalaniTuberculosis (TB), a deadly disease second to HIV/AIDS, is a global health problem. Diagnosis of active tuberculosis is tedious and requires expensive procedures since there is no recognizable method for sole detection of active TB. Although this is a deadly disease, treatment drug toxicity is also an issue that also causes fatalities in diagnosed patients. Therefore, a rapid sensitive and specific diagnostic method is imperative for TB drug management. In this study spectroscopic and/or electrochemical biosensors were developed for the detection and quantification of TB treatment drugs. The biosensors were constructed with electroactive layers of graphene oxide coupled to silver nanoparticles and/or zinc oxide nanoparticles. These nanoparticles coupled with graphene oxide sheets were covalently attached onto the enzymes such as Cytochrome P450-2D6 to achieve the electrochemical detection of the TB treatment drugs and obtain the required electron transfer between the electrode surface and enzyme. The surface morphology of graphene oxide, nanoparticles as well as the green synthesized nanocomposites were achieved using High-Resolution Transmission Electron Microscopy (HRTEM), Atomic Force Microscopy (AFM), and High- Resolution Scanning Electron Microscopy (HRSEM) while the elemental analysis were obtained using Fourier Transform Infrared Spectroscopy (FTIR), Energy Dispersive X-Ray (EDX), Raman spectroscopy and X-Ray diffraction (XRD). Additionally, the optical properties of the developed nanocomposites where further characterised using Small Angle X-ray Scattering (SAXS), Photoluminescence Spectroscopy (PL) and Ultraviolet Spectroscopy (UV-vis). The electrochemical studies were obtained using cyclic voltammetry (CV) and showed an increase in electron conductivity for the green synthesized zinc oxide nanoparticles coupled with graphene oxide (ZnONPs/GO) and silver nanoparticles coupled with graphene oxide (AgNPs/GO) nanocomposite which was an indication that they were suitable as platforms towards biosensor development. Furthermore, amperometric technique was also used for biotransformation of the TB treatment drugs (Ethambutol and Pyrazinamide) in standard solutions of 0.1 M phosphate buffer (pH 7.0). Furthermore, the sensitivity value of 0.0748 μA/μM was determined for the ethambutol biosensor while a value of 0.1715 μA/μM was determined for the pyrazinamide biosensors. Very good detection limits were obtained for the standard solutions of ethambutol and pyrazinamide where a value of 0.02057 nM was determined for ethambutol at concentration linear range of 50 μM – 400 μM. Additionally, a value of 0.8975 x 10-2 nM was determined for pyrazinamide at the concentration linear range of 100 μM – 300 μM. The determined limit of detections have provided a clear indication that these biosensors have potential of being used in human samples since these values are below the peak serum concentrations of these drugs in TB diagnosed patients as reported in literature. This was further confirmed by the limit of quantification values determined for each biosensor where a value of 0.8975 x 10-2 nM was determined for pyrazinamide and a value of 0.02057 nM was determined for ethambutol.