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Study on PHA production by selected thermophillic bacteria
Brondová, Zuzana ; Müllerová, Lucie (referee) ; Obruča, Stanislav (advisor)
This bachelor thesis deals with the study of thermophilic bacterial strains Tepidiphilus thermophilus and Chelatococcus daeguensis aimed at the production of polyhydroxyalkanoates (PHA). Experiments were developed to determine properties of use of these microorganisms in the biotechnological industry. In the experimental parts, the presence of the phaC gene indicating PHA production was confirmed by both bacteriological strains by polymerase chain reaction (PCR). Subsequently, the conditions for PHA production were optimized. Condition profiling aimed at determining the optimal temperature, optimal carbon source, and the effect of the precursor on cell growth, composition and amount of PHA through GC-FID. The last experiment was to determine the utilization of selected substrates. The optimal conditions for the Tepidiphilus thermophilus strain were temperature above 55 ° C and glycerol as the best carbon source. Precursors inhibited cell growth, but the highest amount of 3HV was determined when valeric acid was added to medium. The Chelatococcus daeguensis strain had an optimal temperature of 45 ° C and glucose was optimal carbon source. The best cell metabolizing precursor was 1,4-butanediol. Both bacterial strains were capable of utilizing many substrates where glucose and valeric acid were best.
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Utilization of selected extremophilic bacteria for bioplastic production
Vidláková, Michaela ; Obručová,, Hana (referee) ; Obruča, Stanislav (advisor)
This bachelor thesis is focused on the study of the biotechnological potential of thermophilic bacteria Tepidiphilus thermophilus, Tepidimonas taiwanensis and Tepidimonas fonticaldi. The aim of this work was to investigate the ability of bacteria to produce polyhydroxyalkanoates (PHA). The theoretical part includes the characteristics of PHA and extremophilic bacteria. In the experimental part, the investigated bacteria were tested by molecular technique PCR for the presence of the phaC gene encoding for the first class PHA synthase. The presence of the phaC gene was confirmed in bacterial strains Tepidiphilus thermophilus and Tepidimonas taiwanensis. The ability of PHA production on various carbon sources was examined for all the selected microorganisms, optimal temperatures for cultivation were determined, and the ability to synthesize copolymers was investigated by adding suitable precursors to the production medium. The most intensive PHA production was recorded in the bacterial strain Tepidimonas taiwanensis, but other strains were also able to produce PHA. The ability to produce P(3HB-co-3HV) or P(3HB-co-4HB) copolymers has also been demonstrated in some strains. A significant amount of P(3HB-co-3HV) was obtained using the Tepidimonas taiwanensis strain. Morever, Tepidimonas taiwanensis strain was also tested for its ability to use various model lignocellulose hydrolysates to form PHA. The highest PHA gain was obtained using model hydrolysate of softwood. Furthermore, the ability of proteases production by selected bacterial strains was tested. The presence of these enzymes was detected in the Tepidimonas taiwanensis strain, which was followed by determination of the temperature optima of the proteases, the pH optima of the proteases and also of their molecular weights.
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Characterization of PHA producing microbial cells by advanced microscopic and cytometric techniques
Dlouhá, Karolína ; Nováčková, Ivana (referee) ; Obruča, Stanislav (advisor)
The aim of this bachelor thesis is to document production of polyhydroxyalkanoates in selected bacterial strains, which were Pseudomonas thermotolerans, Chelatococcus daeguensis, Tepidiphilus thermophilus and Chelatococcus thermostellatus. In the case of the microorganisms Tepidiphilus thermophilus and Pseudomonas thermotolerans, the production of PHA´s has not yet been described, and in the Chelatococcus bacteria, which were analysed in this work, the production has not yet been documented by electron microscopy. In this work, the selected producers were analysed by flow cytometry first, using BODIPY and Nile Red as fluorescence probes. Selected producers, for which was production confirmed, were analysed by other methods, such as fluorescence microscopy, cryo scanning electron microscopy and transmission electron microscopy. In the bacterial culture of Pseudomonas thermotolerans, PHA´s production wasn´t confirmed by the first analysis by flow cytometer. For other microorganisms was production confirmed. Chelatococcus bacteria clearly proved to be better producers. Bacterial cells of Tepidiphilus thermophilus produced smaller granules and in lower amount.
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Utilization of selected extremophilic bacteria for bioplastic production
Vidláková, Michaela ; Obručová,, Hana (referee) ; Obruča, Stanislav (advisor)
This bachelor thesis is focused on the study of the biotechnological potential of thermophilic bacteria Tepidiphilus thermophilus, Tepidimonas taiwanensis and Tepidimonas fonticaldi. The aim of this work was to investigate the ability of bacteria to produce polyhydroxyalkanoates (PHA). The theoretical part includes the characteristics of PHA and extremophilic bacteria. In the experimental part, the investigated bacteria were tested by molecular technique PCR for the presence of the phaC gene encoding for the first class PHA synthase. The presence of the phaC gene was confirmed in bacterial strains Tepidiphilus thermophilus and Tepidimonas taiwanensis. The ability of PHA production on various carbon sources was examined for all the selected microorganisms, optimal temperatures for cultivation were determined, and the ability to synthesize copolymers was investigated by adding suitable precursors to the production medium. The most intensive PHA production was recorded in the bacterial strain Tepidimonas taiwanensis, but other strains were also able to produce PHA. The ability to produce P(3HB-co-3HV) or P(3HB-co-4HB) copolymers has also been demonstrated in some strains. A significant amount of P(3HB-co-3HV) was obtained using the Tepidimonas taiwanensis strain. Morever, Tepidimonas taiwanensis strain was also tested for its ability to use various model lignocellulose hydrolysates to form PHA. The highest PHA gain was obtained using model hydrolysate of softwood. Furthermore, the ability of proteases production by selected bacterial strains was tested. The presence of these enzymes was detected in the Tepidimonas taiwanensis strain, which was followed by determination of the temperature optima of the proteases, the pH optima of the proteases and also of their molecular weights.
|
|
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Study on PHA production by selected thermophillic bacteria
Brondová, Zuzana ; Müllerová, Lucie (referee) ; Obruča, Stanislav (advisor)
This bachelor thesis deals with the study of thermophilic bacterial strains Tepidiphilus thermophilus and Chelatococcus daeguensis aimed at the production of polyhydroxyalkanoates (PHA). Experiments were developed to determine properties of use of these microorganisms in the biotechnological industry. In the experimental parts, the presence of the phaC gene indicating PHA production was confirmed by both bacteriological strains by polymerase chain reaction (PCR). Subsequently, the conditions for PHA production were optimized. Condition profiling aimed at determining the optimal temperature, optimal carbon source, and the effect of the precursor on cell growth, composition and amount of PHA through GC-FID. The last experiment was to determine the utilization of selected substrates. The optimal conditions for the Tepidiphilus thermophilus strain were temperature above 55 ° C and glycerol as the best carbon source. Precursors inhibited cell growth, but the highest amount of 3HV was determined when valeric acid was added to medium. The Chelatococcus daeguensis strain had an optimal temperature of 45 ° C and glucose was optimal carbon source. The best cell metabolizing precursor was 1,4-butanediol. Both bacterial strains were capable of utilizing many substrates where glucose and valeric acid were best.
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Characterization of PHA producing microbial cells by advanced microscopic and cytometric techniques
Dlouhá, Karolína ; Nováčková, Ivana (referee) ; Obruča, Stanislav (advisor)
The aim of this bachelor thesis is to document production of polyhydroxyalkanoates in selected bacterial strains, which were Pseudomonas thermotolerans, Chelatococcus daeguensis, Tepidiphilus thermophilus and Chelatococcus thermostellatus. In the case of the microorganisms Tepidiphilus thermophilus and Pseudomonas thermotolerans, the production of PHA´s has not yet been described, and in the Chelatococcus bacteria, which were analysed in this work, the production has not yet been documented by electron microscopy. In this work, the selected producers were analysed by flow cytometry first, using BODIPY and Nile Red as fluorescence probes. Selected producers, for which was production confirmed, were analysed by other methods, such as fluorescence microscopy, cryo scanning electron microscopy and transmission electron microscopy. In the bacterial culture of Pseudomonas thermotolerans, PHA´s production wasn´t confirmed by the first analysis by flow cytometer. For other microorganisms was production confirmed. Chelatococcus bacteria clearly proved to be better producers. Bacterial cells of Tepidiphilus thermophilus produced smaller granules and in lower amount.
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