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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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Pregnancy proteins - molecular biological and biochemical analysis
Muravská, Alexandra ; Kalousová, Marta (advisor) ; Černá, Marie (referee) ; Průša, Richard (referee)
The aim of this thesis was to establish methods for selected PAPP-A (Pregnancy- Associated Plasma Protein A) gene polymorphisms analysis and to study genetic background of PAPP-A and biochemical background of PAPP-A and PlGF (Placental Growth Factor) in relation to risk pregnancy. Secondly, the aim was to establish method for two-dimensional (2D) electrophoresis of amniotic fluid. Methods for analysis of ten PAPP-A gene polymorphisms were established. These polymorphisms, PAPP-A and PlGF levels were studied in together 165 women in third trimester pregnancies complicated with threatening preterm labor (n=98), preeclampsia (n=35), IUGR (Intrauterine Growth Restriction) (n=34) and ICP (Intrahepatic Cholestasis of Pregnancy) (n=15). 114 healthy pregnant women served as controls. The method for 2D electrophoresis of amniotic fluid was established. Preeclamptic patients had significantly higher frequency of TT genotype of Cys327Cys (C/T) PAPP-A gene polymorphism compared to controls. Patients with ICP had increased serum levels of PAPP-A compared to controls, in patients with threatening preterm labor PAPP-A levels were rather decreased. PlGF levels did not differ from control group in patients with ICP and threatening preterm labor. Positive correlation was found between PAPP-A and PlGF in group of...
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Signal processing based methods for genome assembly refinement
Jugas, Robin ; Provazník, Ivo (referee) ; Sedlář, Karel (advisor)
The diploma thesis deals with sequencing methods and genome assembly methods including usage of numerical representations. The theoretical part of thesis describes the history of DNA research, generations of sequencing methods, the assembly methods themselves and definiton of numerical representations. Numerical represenatations serve to convert character form of DNA to numerical form and so allow to use digital signal processing methods. There is an algorithm for genome assembly using numerical represenatation proposed in thesis, which is later tested at sequence data.
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Genotyping of bacterial genus \kur{Xanthomonas} pathogenic for tomato and pepper by multilocus sequencing.
STEHLÍKOVÁ, Dagmar
The subject of this bachelor thesis is to estimate the degree of relationship of 20 collected isolated samples of Xanthomonas genus causing bacterial spot of tomatoes and peppers. To assess the relationship, the method of multilocus sequence analysis has been used (MLSA). Sequences of genes atpD, rpoD, gyrB, glnA and efP were found and analyzed using MEGA software version 5.1. The result was evaluated using a dendrogram according to which the isolates were divided into two clusters. The first cluster includes isolated of Xanthomonas euvesicatoria belonging to group A. The second cluster includes isolates of the species Xanthomonas vesicatoria belonging to group B. The results obtained in this work are consistent with the present classification of the xanhtomonads causing bacterial spot of tomatoes and peppers.
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