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Evaluation of a micromethod for isolation of DNA from plant leaf, fruit and fruit products
Balažovičová, Nikola ; Rittich, Bohuslav (referee) ; Kovařík, Aleš (advisor)
The thesis has been focused on testing of micromethod of DNA isolation from leaves, fruits and fruit products. Jams were selected for the analysis of plant DNA in technologically processed foods. Plant leaves, fruits, and jams were homogenized using plastic copist in a lysis buffer containing 2% cetyltrimethylammonium bromide (CTAB) with 2.5M sodium chloride (NaCl). Microisolation of plant DNA was performed using poly(hydroxyethylmethacrylate-co-glycidylmethacrylate) – P(HEMA-co-GMA)microparticles. Isolated the DNA concentration and purity were assessed by UV light aborbance using a spectrophotometer. After that, amplification of the DNA was tested in PCR. Primers specific for plant ribosomal DNA: 18S_for a 5,8S_rev (PCR product - 700bp), 26S_for a 26S_rev (PCR product - 220 bp), 18S_for a 18¬S_rev (PCR product - 263 bp) were used. The PCR conditions were optimized and the effect of the amplicon length on its detection was followed. PCR products were detected by agarose gel electrophoresis. It was shown that DNA isolated from almost all of leaves using magnetic particles was in PCR-ready quality in contrary to the fruits. DNA amplified in PCR with primers giving short PCR products was isolated from almost all tested jams. The method must be optimalised, yet.
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The safety of probiotics used in foods
Balažovičová, Nikola ; Rittich, Bohuslav (referee) ; Španová, Alena (advisor)
The theoretical part of this thesis focuses on definition of probiotics, their health benefits, probiotic foods, safety criteria for probiotics used in foods, including methods for the identification of probiotic bacteria. Experimental part of work was focused on the identification of probiotic bacteria contained in the milk product Actimel. DNA quality suitable for the PCR was isolated by magnetic microparticles P(HEMA-co-GMA). Identification was estimated using the method of polymerase chain reaction. The presence of DNA of domain Bacteria, genus Lactobacillus and species Lactobacillus casei was confirmed by gel electrophoresis of PCR products. Specific PCR products of the sizes of 466 bp (domain Bacteria), 250 bp (genus Lactobacillus) and 136 bp (Lactobacillus casei) were detected after amplification of DNA isolated from a probiotic milk product.
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Evaluation of a micromethod for isolation of DNA from plant leaf, fruit and fruit products
Balažovičová, Nikola ; Rittich, Bohuslav (referee) ; Kovařík, Aleš (advisor)
The thesis has been focused on testing of micromethod of DNA isolation from leaves, fruits and fruit products. Jams were selected for the analysis of plant DNA in technologically processed foods. Plant leaves, fruits, and jams were homogenized using plastic copist in a lysis buffer containing 2% cetyltrimethylammonium bromide (CTAB) with 2.5M sodium chloride (NaCl). Microisolation of plant DNA was performed using poly(hydroxyethylmethacrylate-co-glycidylmethacrylate) – P(HEMA-co-GMA)microparticles. Isolated the DNA concentration and purity were assessed by UV light aborbance using a spectrophotometer. After that, amplification of the DNA was tested in PCR. Primers specific for plant ribosomal DNA: 18S_for a 5,8S_rev (PCR product - 700bp), 26S_for a 26S_rev (PCR product - 220 bp), 18S_for a 18¬S_rev (PCR product - 263 bp) were used. The PCR conditions were optimized and the effect of the amplicon length on its detection was followed. PCR products were detected by agarose gel electrophoresis. It was shown that DNA isolated from almost all of leaves using magnetic particles was in PCR-ready quality in contrary to the fruits. DNA amplified in PCR with primers giving short PCR products was isolated from almost all tested jams. The method must be optimalised, yet.
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|
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The safety of probiotics used in foods
Balažovičová, Nikola ; Rittich, Bohuslav (referee) ; Španová, Alena (advisor)
The theoretical part of this thesis focuses on definition of probiotics, their health benefits, probiotic foods, safety criteria for probiotics used in foods, including methods for the identification of probiotic bacteria. Experimental part of work was focused on the identification of probiotic bacteria contained in the milk product Actimel. DNA quality suitable for the PCR was isolated by magnetic microparticles P(HEMA-co-GMA). Identification was estimated using the method of polymerase chain reaction. The presence of DNA of domain Bacteria, genus Lactobacillus and species Lactobacillus casei was confirmed by gel electrophoresis of PCR products. Specific PCR products of the sizes of 466 bp (domain Bacteria), 250 bp (genus Lactobacillus) and 136 bp (Lactobacillus casei) were detected after amplification of DNA isolated from a probiotic milk product.
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