Browsing by Author "Rees, Jasper G."
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Item Alteration of Bax-to-Bcl-2 ratio modulates the anticancer activity of methanolic extract of Commelina benghalensis (Commelinaceae) in Jurkat T cells(Academic Journals, 2008) Mbazima, Vusi G.; Mokgotho, Matlou P.; February, Faghri; Rees, Jasper G.; Mampuru, Leseilane J.Stem extracts of Commelina benghalensis (Linn.), although not extensively documented, are frequently used in traditional medicine for the treatment of ailments such as skin malformations and outgrowths. Accordingly, the study was aimed to investigate possible molecular mechanisms that are associated with the potential anti-carcinogenic property of this agrofield weed. Jurkat T cells were exposed to different concentrations (0-600 ug/ml) of the crude methanolic extract of C. benghalensis to evaluate their growth inhibitory and apoptosis inducing effects. The extract elicited a dose- and time-dependent inhibition of cell proliferation, followed by a concomitant decrease in cell viability. The observed cytotoxicity was linked to the induction of apoptosis as determined by morphological and biochemical features known to be associated with the advent of apoptosis. Real time quantitative RT-PCR and Western blot analyses of Bax, Bcl-2 and p53 exhibited aberrant expression profiles of these genes under various treatment conditions. Taken together, the data suggest that the crude methanolic extract of C. benghalensis contains bioactive compounds that may be beneficial in the treatment of malignant growths, and that this apparent antineoplastic activity is a consequence of dysregulated expression of apoptosis-responsive genes. These observations could provide a credible scientific justification upon which the ethnopharmacological utilisation of C. benghalensis is founded.Item Genome-wide SNP identification by high-throughput sequencing and selective mapping allows sequence assembly positioning using a framework genetic linkage map(BioMed Central, 2010) Celton, Jean M.; Christoffels, Alan; Sargant, Daniel J.; Xu, Xiangming; Rees, Jasper G.Determining the position and order of contigs and scaffolds from a genome assembly within an organism’s genome remains a technical challenge in a majority of sequencing projects. In order to exploit contemporary technologies for DNA sequencing. We developed a strategy for whole genome single nucleotide polymorphism sequencing allowing the positioning of sequence contigs onto a linkage map using the bin mapping method. The strategy was tested on a draft genome of the fungal pathogen Venturia inaequalis, the causal agent of apple scab, and further validated using sequence contigs derived from the diploid plant genome Fragaria vesca. Using our novel method we were able to anchor 70% and 92% of sequences assemblies for V. inaequalis and F. vesca, respectively, to genetic linkage maps. We demonstrated the utility of this approach by accurately determining the bin map positions of the majority of the large sequence contigs from each genome sequence and validated our method by mapping single sequence repeat markers derived from sequence contigs on a full mapping population.Item A glance at quality score: implication for de novo transcriptome reconstruction of Illumina reads(Frontiers, 2014) Mbandi, Stanley K.; Hesse, Uljana; Rees, Jasper G.; Christoffels, AlanDownstream analyses of short-reads from next-generation sequencing platforms are often preceded by a pre-processing step that removes uncalled and wrongly called bases. Standard approaches rely on their associated base quality scores to retain the read or a portion of it when the score is above a predefined threshold. It is difficult to differentiate sequencing error from biological variation without a reference using a quality score. The effects of quality score based trimming have not been systematically studied in de novo transcriptome assembly. Using RNA-Seq data produced from Illumina,we teased out the effects of quality score based filtering or trimming on de novo transcriptome reconstruction. We showed that assemblies produced from reads subjected to different quality score thresholds contain truncated and missing transfrags when compared to those from untrimmed reads. Our data supports the fact that de novo assembling of untrimmed data is challenging for de Bruijn graph assemblers. However, our results indicates that comparing the assemblies from untrimmed and trimmed read subsets can suggest appropriate filtering parameters and enables election of the optimum de novo transcriptome assembly in non-model organisms.Item Selecting an appropriate method for expressing S locus F-box-S2 recombinant protein(Elsevier, 2017) Ashkani, Jahanshah; Rees, Jasper G.A single locus (S locus) including at least two linked genes (female and male determinants) genetically controls the gametophytic self-incompatibility (GSI) in apple, which has evolved to avoid self-fertilization. There has been extensive work done on the female determinant of self-incompatibility, which has led to the determination of the tertiary structure of S-RNase. However, the tertiary structure of male determinant (S locus F-box, SLF/SFB) remains unresolved, which could mainly be due to difficulties associated with its expression in the recombinant expression systems. In addressing this, we have evaluated several in vivo (prokaryotic and eukaryotic) and in vitro expression systems for their efficiency in the expression of apple SLF2. The most successful expression of SLF2 (1 mg/ml) was achieved in E. coli using the synthesized gene in a high salt culture and applying heat shock before induction of culture. We therefore present an approach for the efficient expression of S locus F-box recombinant proteins for future functional and structural studies.