Molecular investigation of stalk formation and growth rates in Didymosphenia geminata

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2018-04-02
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Abstract
Didymosphenia geminata, a freshwater diatom, can produce thick benthic mats of stalk material. The cause of excess stalk formation is proposed to be low soluble reactive phosphorus (SRP) concentrations or a mat-forming genetic variant. Molecular studies on D. geminata are few, partly due to the lack of a reliable DNA extraction technique for the diatom, but have the ability to elucidate the drivers of mat formation. Specifically, comparison of gene expression between mat and non-mat-forming sites using reference genomes, may reveal underlying mechanisms behind growth and stalk formation. The objectives of this study were to: 1) identify gene products, expressed only at a mat or non-mat-forming site in Alberta (AB), involved in cell division or stalk formation from the reference genomes of P. tricornutum, T. pseudonana and C. crescentus; 2) compare SRP and growth rate, using frequency of dividing cells (FDC), between a mat-forming and non-mat-forming site; 3) compare FDC between multiple mat-forming sites in AB and British Columbia (BC); and 4) compare the quantity and quality of DNA obtained from mat-forming D. geminata using various preservation buffers, storage temperatures and extraction methods, including organic extraction, the Qiagen DNeasy® Plant Mini Kit, and a version of the kit procedure modified for D. geminata. Sixty-six putative gene products from the reference genomes were expressed only in the mat-forming sample, while 172 were expressed only in the non-mat-forming sample. FDC and SRP were higher at the non-mat-forming site, with no significant difference between FDC in AB and BC. Collectively, these results support previous findings that D. geminata forms excess stalk material under low SRP and increases cell division under higher SRP. However, an anomalous observation of mat formation at a site with 0.00986 mg/L SRP indicates stalk formation may be influenced by additional factors. In addition, the results of this study indicate that DNA extracted from samples stored in 95% EtOH at -80°C, using the modified kit method, had the best combination of quantity and quality.
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Larsen, K. M. (2018). Molecular investigation of stalk formation and growth rates in Didymosphenia geminata (Master's thesis, University of Calgary, Calgary, Canada). Retrieved from https://prism.ucalgary.ca. doi:10.11575/PRISM/31764