Browsing by Author "Gupthar, Abindra Supersad."

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Analyses of lipidic bodies from green microalgae.
(2014) Pather, Verushka.; Gupthar, Abindra Supersad.; Bux, Faizal.
This study presents the analyses of oil body components in microalgae which may be involved in oil droplet assembly including certain triacylglycerol precursors which can be processed to biodiesel, an alternative fuel source. Stress induction of microalgae, Chlorella vulgaris CCAP 211/11B and Dunaliella primolecta CCAP 11/34 was achieved by exclusion of nitrates in growth media. Contrary to other forms of nitrogen depletion, this condition did not greatly enhance lipid biosynthesis in the microalgae. Confocal microscopy and fluorescent dyes nile red and bodipy were employed for the visualization of lipidic body components. The fluorescence hues emitted by neutral lipids and phospholipids were differentiated from those due to autofluorescence and chlorophyll using ZEN software to analyse images from a Zeiss LSM 710 confocal microscope. Oil from both algae, which were subjected to transesterification and gas chromatography, revealed a predominant fatty acid, namely palmitic acid (C16:0). D. primolecta produced linolelaidic acid (C18:2n6t) under growth conditions involving both nitrate supplementation and exclusion; whilst the longest fatty acid, docosanoic acid (C22:0 chain) was produced by the alga C. vulgaris only under conditions of nitrate supplementation. Nitrate limitation had minimal effect on the oil hydrocarbon yield which increased only 0.02% and 0.01% for C. vulgaris and D. primolecta, respectively. The highest biodiesel yield of 26.11 % was recorded from D. primolecta when grown under conditions of nitrate exclusion. The protein concentrations extracted from oil of the former alga ranged from 1.87 - 1.95 Gg/ml when grown under nitrate supplemented conditions and 1.74 - 1.90 Gg/ml when nitrate was excluded from the media. The protein concentrations extracted from oil of D. primolecta ranged from 1.91 - 2.23 Gg/ml and 1.88 - 1.98 Gg/ml, respectively, when the algae were grown in the presence and exclusion of nitrates. In the adaptation of protocols for protein extraction from oil, sunflower and salmon oils were initially used. Sunflower oil extracts produced by 10% (w/v) SDS treatment, yielded protein bands of 198, 96, 70 and 58 KDa on 10% (w/v) polyacrylamide gels while 6M urea treatment yielded a band of 200 KDa. Salmon oil treated with 10% (w/v) SDS and 6 M urea yielded bands of 195 and 27 KDa, and 198 KDa, respectively, as well as common bands of 68 and 64 KDa. In comparison, the extraction of discrete proteins from algal oil proved to be difficult as the extractants SDS and urea could have denatured protein components into subunit structure.
Biochemistry students' difficulties with the symbolic and visual language used in molecular biology.
(2007) Gupthar, Abindra Supersad.; Anderson, Trevor Ryan.
This study reports on recurring difficulties experienced by undergraduate students with respect to understanding and interpretation of certain symbolism, nomenclature, terminology, shorthand notation, models and other visual representations employed in the field of Molecular Biology to communicate information. Based on teaching experience and guidelines set out by a four-level methodological framework, data on various topic-related difficulties was obtained by inductive analyses of students’ written responses to specifically designed, free-response and focused probes. In addition, interviews, think-aloud exercises and student-generated diagrams were also used to collect information. Both unanticipated and recurring difficulties were compared with scientifically correct propositional knowledge, categorized and subsequently classified. Students were adept at providing the meaning of the symbol “Δ” in various scientific contexts; however, some failed to recognize its use to depict the deletion of a leucine biosynthesis gene in the form, Δ leu. “Hazard to leucine”, “change to leucine” and “abbreviation for isoleucine” were some of the erroneous interpretations of this polysemic symbol. Investigations on these definitions suggest a constructivist approach to knowledge construction and the inappropriate transfer of knowledge from prior mental schemata. The symbol, “::”, was poorly differentiated by students in its use to indicate gene integration or transposition and in tandem gene fusion. Idiosyncratic perceptions emerged suggesting that it is, for example, a proteinaceous component linking genes in a chromosome or the centromere itself associated with the mitotic spindle or “electrons” between genes in the same way that it is symbolically shown in Lewis dot diagrams which illustrate covalent bonding between atoms. In an oligonucleotide shorthand notation, some students used valency to differentiate the phosphite trivalent form of the phosphorus atom from the pentavalent phosphodiester group, yet the concept of valency was poorly understood. By virtue of the visual form of a shorthand notation of the 3,5 phosphodiester link in DNA, the valency was incorrectly read. VSEPR theory and the Octet Rule were misunderstood or forgotten when trying to explain the valency of the phosphorus atom in synthetic oligonucleotide intermediates. Plasmid functional domains were generally well-understood although restriction mapping appeared to be a cognitively demanding task. Rote learning and substitution of definitions were evident in the explanation of promoter and operator functions. The concept of gene expression posed difficulties to many students who believed that genes contain the entity they encode. Transcription and translation of in tandem gene fusions were poorly explained by some students as was the effect of plasmid conformation on transformation and gene expression. With regard to the selection of transformants or the hybridoma, some students could not engage in reasoning or lateral thinking as protoconcepts and domain-specific information were poorly understood. A failure to integrate and reason with factual information on phenotypic traits, media components and biochemical pathways were evident in written and oral presentations. DNA-strand nomenclature and associated function were problematic to some students as they failed to differentiate coding strand from template strand and were prone to interchange the labelling of these. A substitution of labels with those characterizing DNA replication intermediates demonstrated erroneous information transfer. DNA replication models posed difficulties integrating molecular mechanisms and detail with line drawings, coupled with inaccurate illustrations of sequential replication features. Finally, a remediation model is presented, demonstrating a shift in assessment score dispersion from a range of 0 - 4.5 to 4 - 9 when learners are guided metacognitively to work with domain-specific or critical knowledge from an information bank. The present work shows that varied forms of symbolism can present students with complex learning difficulties as the underlying information depicted by these is understood in a superficial way. It is imperative that future studies be focused on the standardization of symbol use, perhaps governed by convention that determines the manner in which threshold information is disseminated on symbol use, coupled by innovative teaching strategies which facilitate an improved understanding of the use of symbolic representations in Molecular Biology. As Molecular Biology advances, it is likely that experts will continue to use new and diverse forms of symbolic representations to explain their findings. The explanation of futuristic Science is likely to develop a symbolic language that will impose great teaching challenges and unimaginable learning difficulties to new generation teachers and learners, respectively.
Constitutive over-expression of FL01 encoded mannoproteins.
(2014) Ngobese, Lethukuthula Melusi.; Govender, Patrick.; Gupthar, Abindra Supersad.
The ability of yeast cell wall-derived mannoproteins (glycoproteins) to positively contribute to oenological properties has been a key factor that stimulates research initiatives into these industrially important glycoproteins. In addition, and from a fundamental research perspective, yeast cell wall glycoproteins are involved in a wide range of biological interactions. To date, and to the best of our knowledge, biochemical insight into the fine molecular structure of these mannoproteins is fairly limited. The amino acid sequences of their protein moieties have been established from structural and functional in silico analysis of the genomic sequence of Saccharomyces cerevisiae whilst far less structural details are available on the glycosyl moieties of these mannoproteins. In an attempt to generate significant quantities of cell wall-attached glycoproteins that may be employed for the purpose of biochemical structural analysis, a novel strategy was envisaged that encompasses the genetic manipulation of S. cerevisiae strains to over-express and release cell wall-associated glycoproteins into the liquid growth medium. To this end, the FLO1-encoded cell wall mannoprotein was overexpressed in haploid S. cerevisiae laboratory strains bearing a specific cell wall biosynthesis-related gene deletion that has been previously shown to promote extracellular hyper-secretion of cell wall-based glycoproteins that would otherwise be attached to the cell wall in the feral strain. A polymerase chain reaction (PCR)-based cloning stratagem was employed to yield transgenic strains in which the native FLO1 open reading frame (ORF) is placed under transcriptional control of the constitutive PGK1 promoter. The data reveals that the strategy employed resulted in the expected constitutive expression pattern of the FLO1 ORF in all transgenic strains created in this study. A modified Helm’s flocculation assay was employed to assess flocculation intensities of the three FLO1-based transgenic strains. This assay confirmed that the flocculation phenotypes observed were indeed of the Flo1-type in that they were Ca²⁺-dependent, sensitive to ethylenediaminetetraacetic acid (EDTA) and inhibited in the presence of mannose. Interestingly, the KNR4 deletion-based transgenic yeast strain displayed a visibly weaker different adhesion behaviour in terms of weaker flocculation intensity as evidenced by noticeably smaller floc aggregate size. In addition a greater quantity of proteins was detected at statistically significant levels in the spent culture medium of the KNR4 deletion-based transgenic yeast strain. Although yet to be purposefully optimised, the data seemingly suggests that PGK1 promoter-based mediated overexpression of the FLO1 ORF in a KNR4 deletion-based genetic background has the potential to release the desired mannoprotein into the culture medium.
Mathematical modelling of the dynamical interactions between killer and sensitive wine yeast subjected to nutritional stress.
(2000) Vadasz, Alisa S.; Abashar, Mohamed Elbashir Elamin.; Gupthar, Abindra Supersad.
No abstract available.
Spatio-temporal dynamics of heterogeneously distributed populations.
(2003) Vadasz, Alisa S.; Carsky, Milan.; Gupthar, Abindra Supersad.
No abstract available.