Application of molecular method for the detection of freshwater microalgae
| Autor: | Chih-Chiang Hsu, 許智強 |
|---|---|
| Rok vydání: | 2011 |
| Druh dokumentu: | 學位論文 ; thesis |
| Popis: | 99 Algae are the most important primary producers in the aquatic ecosystems and could be found in different habitats with complex phylogenetic diversity. Once the aquatic environment was polluted, indigenous algae community changed correspondently. This effect might not just a reduction in the number of species, but rather a change in the species composition of the community. Therefore monitoring the changes in population of algae become an important issue. Conventional approaches use microscopy to identify the species and numbers of algae. However, microscopic identification is not only time-consuming but also being biased by individual experiences. Furthermore, among different algae species it is not unusual to find very similar shapes, and phenotype-based species often mask genetic diversity as well. Molecular methods is a useful tool for studying algae communities in freshwater. Several recent studies have demonstrated the effectiveness of molecular methods to estimate the diversity of natural prokaryotic communities in aquatic environments. They demonstrated that molecular methods are not only fast and simple but also highly sensitive for the detection and identification of algae. The aims of this study is to compare the community difference between conventional approach (microscopic examination) and molecular methods in attempt to develop a reliable and faster procedure for the detection and quantitative determination of algae in environmental samples. Performance of PCR-DGGE method on exploring the algae diversity was examined in this study. Real-Time PCR method targeting universal 23S plastid rDNA genes was also applied for quantification means. The difference between molecular-based methods (PCR-DGGE, Real-Time PCR) and microscopic analysis were discussed. Furthermore, a set of diatom specific primer targeting 18S rDNA was applied for exploring the communities of diatom. The microscopic examination results showed that the algal communities were dominated by diatoms. However, completely different results were obtained by the 23S plastid rDNA-based PCR-DGGE method in which the algae community were dominated by Synechococcus sp. PCR-DGGE analysis cannot detect any diatoms in all sampling points using this primer set. No Synechococcus sp. could be identified by microscopic examination. We found that there were not in total agreement with the results from Real-Time PCR quantification and cell counting by microscopy. On the other hand, when the diatom specific primer was applied to assess the diatoms community composition, satisfied results were obtained in which the major diatom groups can be identified by PCR-DGGE method. The Real-Time PCR quantification results also presented more correlation with the microscopic analysis on the diatom issus. |
| Databáze: | Networked Digital Library of Theses & Dissertations |
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