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Methods for storing and archiving zoological tissue collections, and identification based on DNA barcoding
Aghová, Tatiana ; Benda, Petr ; Brejcha, Jindřich ; Dolejš, Petr ; Kyralová, Eva ; Mlíkovský, Jiří ; Moravec, Jiří ; Šanda, Radek ; Štundl, Jan ; Tkoč, Michal ; Vondráček, Dominik
Fulltext:  PDF; PDF; PDF; PDF
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CNV detection in bacterial genomes
Lacinová, Michaela ; Sedlář, Karel (referee) ; Škutková, Helena (advisor)
This master thesis deals with analysis of structural variation of genome and with methods of its sequencing across all generations. Subsequently it contains a description of copy number variation and methods of its detection. The experimental part focuses on algorithm proposal for CNV detection according analysis and testing of uneven coverage in genome, variable representation of GC content and distance of sequence reads. Finally, the algorithm for detecting copy number variation is tested on genomic data of bacteria Klebsiella pneumoniae.
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Complex characterization of subgingival plaque - use of modern sequencing methods in diagnostics and monitoring of treatment of periodontal diseases
Těšínská, Barbora ; Najmanová, Lucie (advisor) ; Konopásek, Ivo (referee)
Periodontitis is a multifactorial inflammatory disease which can result in a complete loss of teeth. Its main cause is the accumulation of bacteria from the dental plaque followed by massive reaction of the host immune system. It was proved that the composition of oral microbiome (OM) differs in periodontally healthy individuals and patients with periodontitis. This work aims to solve specific parts of the long-term project concerning the taxonomic composition of the OM of periodontally healthy individuals and patients with chronic and aggressive form of the disease. The OM was characterized based on 16s rDNA sequencing. It is evident from the results that the shift in the OM composition occurs prior the development of clincal signs of the disease and that the precise dental care can significantly postpone or even avoid the onset of the disease. When comparing the OM composition in individuals with chronic and aggressive periodontitis, no remarkable differences were detected to explain the faster progress of the aggressive form of the disease. This work also aimed to compare the results obtained by 454 pyrosequencing and Illumina. Both sequencing methods were found to provide statistically comparable results. Illumina MiSeq thus can be employed to build on the former results of the long-term study...
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Fragment analysis represents a suitable approach for the detection of hotspot c.7541_7542delCT NOTCH1 mutation in chronic lymphocytic leukemia
VONKOVÁ, Barbara
Chronic lymphocytic leukemia is the most common type of leukemia in adults in western countries. Despite the continual improvement of diagnostic methods and treatment, it still remains incurable. One of the main characteristic features of CLL is its clinical and biological variability. In present, the analysis of prognostic genetic markers is the most useful tool for understanding biology and clinical course of the disease and consequently the individual type of treatment. The presence of a NOTCH1 mutation is connected with shortened survival and resistance to conventional chemo-immunotherapy. In this thesis, we examined CLL patients with c.7541_7542delCT NOTCH1 mutation and the most sensitive and specific method was identified. We compared nowadays most commonly used methods: allele-specific PCR, fragment analysis and Sanger sequencing.
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Taxonomic classification of yeasts associated with meadow plants
Čurillová, Natália ; Ing.Hana Dudášová, Ph.D. (referee) ; Horváthová, Ágnes (advisor)
In total 60 yeast strains isolated from meadow flowers were selected for identification using MALDI-TOF biotyping. Selected yeast strains were prepared according to standard (Bruker) method and method developed at Institute of Chemistry SAS in Bratislava for MALDI-TOF yeast identification. To acquire valuable data two cultivating media were used. The principle of identification embody in comparing obtained mass spectra according to the score of selected yeast strain isolated from meadow plants, formerly identified by physicobiochemical properties (verification of their taxonomic classification) or the new isolates (their identification) with mass spectra of the reference yeast cultures. Reference yeast cultures were identified by sequencing the D1/D2 of 26S rRNA domain. In total 53 yeast strains were identified by biotyping at species level. The remaining 7 yeast strains were designed to be identified by sequencing the D1/D2 26S rRNA domain as the reference mass spectra were not available. The highest abundance of yeast species was as follows Metschnikowia pulcherrima (12 %), Rhodotorula spp. (12 %), Cystofilobasidium infirmominiatum (10 %), Metschnikowia reukaufii (7 %), Metschnikowia fructicola (7 %), Sporobolomyces metaroseus (7 %), Hanseniospora uvarum (5 %), Rhodotorula mucilaginosa (5 %), Meyerozyma guillermondii (5 %), Cystofilobasidium macerans (3 %) a Rhodotorula dairenensis (3 %), and yeast strains of species Candida bombi, Pichia kudravzievii, Cystofilobasidium capitatum, Cystofilobasidium macerans, Solicoccozyma aeria, Sporidiobolus salmonicolor, Papiliotrema flavescens, Sporidiobolus metaroseus, Sporidiobolus pararoseus and Cryptococcus wieringae were identified as well. In general, the most diverse abundance of yeast species were isolated from meadow plants comparing to the older studies.
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Verification of somatic mutations important for neuroectodermal tumors (CTNNB1, BRAF, ALK)
Hrindová, Božidara ; Vícha, Aleš (advisor) ; Janatová, Markéta (referee)
The diploma thesis was focused on evidence of selected somatic mutations in genes ALK (Anaplastic lymphoma receptor tyrosine kinase), BRAF (v-Raf murine sarcoma viral oncogene homolog B1) and β-catenin (CTNNB1) through molecular - genetic methods in the target group of neuroectodermal tumors (neuroblastoma, medulloblastoma, brain tumors, paraganglioma and pheochromocytoma). Some of them are already considered as prognostic indicators which help to identify the subtype of various tumors and on the basis of this molecular - biological classification choosing the appropriate treatment. The genetic material of 133 patients was used for the analysis divided by the type of cancer. The presence of the mutation was detected in seven cases, of which two of them beloged to the gene BRAF, one to the gene ALK and four to the gene β-catenin. The subject of research in the cases of this genes were hotspot mutation sites. The purpose was to confirm the presence of the mutation in the hotspots and contribute to the studies which are aimed at the introduction of more suitable treatment through the inhibitors of mutated genes. Keywords: ALK, BRAF, β-catenin (CTNNB1), neuroectodermal tumors, sequencing, MLPA
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Analysis of gene expresion via next generation sequencing techniques
Bláhová, Monika ; Krylov, Vladimír (advisor) ; Žárský, Vojtěch (referee)
1 Abstract The main task of this work is create review today's available methods for gene expression analysis, introduce advantages and disadvantages of this metods and compare them. Nowadays sequencing is one of the most usable molecular methods. Sequencing methods are divided to three groups, a first generation sequencing, a second generation sequencing and a third generation sequencing. The most useful are the second generation sequencing. However, the third generation sequencing have a big potencial too. It is not necessary amplificate samples using PCR thanks them. Amount of data raises rapidly, thanks decreasing costs and increasing efficiency. Demands on data starage, computers output are growing. And softwares for data analysis are much clever than ever before.
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Contribution of Next Generation Sequencing for Laboratory Diagnostics
Votýpka, Pavel ; Beránek, Martin (advisor) ; Macháček, Miloslav (referee)
5 ABSTRACT Charles University in Prague Faculty of Pharmacy in Hradec Králové Department of Biochemical Sciences Candidate: Bc. Pavel Votýpka Supervisor: Doc. PharmDr. Martin Beránek, Ph.D. Consultant: Mgr. Nikola Ptáková Title of diploma thesis: Contribution of Next Generation Sequencing for Laboratory Diagnostics The endeavor to sequence the whole human genome lead not only to the knowledge acquisition regarding the human genetic information but as well to the development of new sequencing methods and technologies. In order to keep up with progress in genetic field in many clinical and research laboratories the new massive parallel sequencing equipment is being utilized. On the market are currently established four leading platforms - Illumina, Solid, Ion Torrent and 454 Life Technologies. The process of sequencing analysis can be summarized into three main steps - the sequencing library preparation, sequencing itself, variant calling and data analysis. Each part of the sequencing analysis exhibits certain specifics, we need to count with and as well its pitfalls, we need to avoid or to minimalize their impact on the analysis final result. Recently new methods termed sequencing of the 3rd generation are being developed, enabling sequence of a single DNA molecule to be determined without previous...
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The use of environmental sequencing in the studies on eukaryotic diversity
Lukešová, Soňa ; Hampl, Vladimír (advisor) ; Škaloud, Pavel (referee)
The bachelor's thesis focuses on environmental sequencing method and its usage in examining the diversity of microbial eukaryotic organisms. It describes the method, its mechanism, application and the problems associated with it. The work describes the current view of the phylogeny of the major eukaryotic lineages with their closer characteristics. Particular attention is paid to newly discovered groups. The work addresses the relationships inside these groups, summarizes findings of their ecology, food resources, and morphology. A separate chapter is devoted to the occurrence of the protist groups in extreme environmental conditions. Key words: Diversity of eukaryotes, environmental DNA, sequencing, rRNA, phylogenetics.
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