|
|
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.
|
|
|
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.
|
|
| |
|
|
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
|
|
|
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.
|
|
|
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...
|
|
|
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.
|
|
| |
|
|
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...
|
|
|
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.
|
guest ::
