Browsing by Author "Maropola, Mapula Kgomotso Annah"

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    Identification of endophytic bacterial communities associated with South African crops: sorghum bicolor (L. Moench), pennisetum glaucum and arachis villosulicarpa
    (University of the Western Cape, 2014) Maropola, Mapula Kgomotso Annah; Tuffin, Marla
    In this study, the diversity of endophytic bacteria associated with food crops, sorghum (Sorghum bicolor L. Moench), pearl millet (Pennisetum glaucum L.) and groundnut (Arachis villosulicarpa) is investigated using culture-independent techniques: terminal retriction fragment length polymorphism (t- RFLP) and next generation sequencing (NGS). The first objective of this study was to investigate the effect of different DNA extraction protocols on mDNA yield and quality, as well as the diversity of endophytic bacteria retrieved from root and stem tissues (0.1g or 0.3g) of sorghum, pearl millet and groundnut. Protocols used include two classical methods (CTAB- and SDS-based) and five commercial kits: MoBio PowerPlant Pro® DNA Isolation Kit, Qiagen DNeasyR Plant Mini Kit, Fermentas GeneJET Plant Genomic DNA Purification Kit, MoBio PowerSoilTM DNA Purification Kit and MoBio UltraClean® Soil DNA Isolation Kit. Eletrophoresis and the Nanodrop were used to determine DNA yield and purity
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    Impact of metagenomic DNA extraction procedures on the identifiable endophytic bacterial diversity in Sorghum bicolor (L. Moench)
    (Elsevier, 2015) Maropola, Mapula Kgomotso Annah; Ramond, Jean-Baptiste; Trindade, Marla
    Culture-independent studies rely on the quantity and quality of the extracted environmental metagenomic DNA (mDNA). To fully access the plant tissue microbiome, the extracted plant mDNA should allow optimal PCR applications and the genetic content must be representative of the total microbial diversity. In this study, we evaluated the endophytic bacterial diversity retrieved using different mDNA extraction procedures. Metagenomic DNA from sorghum (Sorghum bicolor L. Moench) stem and root tissues were extracted using two classical DNA extraction protocols (CTAB- and SDS-based) and five commercial kits. The mDNA yields and quality as well as the reproducibility were compared. 16S rRNA gene terminal restriction fragment length polymorphism (t-RFLP) was used to assess the impact on endophytic bacterial community structures observed. Generally, the classical protocols obtained high mDNA yields from sorghum tissues; however, they were less reproducible than the commercial kits. Commercial kits retrieved higher quality mDNA, but with lower endophytic bacterial diversities compared to classical protocols.