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DNA topology as a factor affecting transcription in bacteria
Hladíková, Kristýna ; Krásný, Libor (advisor) ; Nešvera, Jan (referee)
DNA topology plays a key role in regulation of gene expression by affecting interactions between DNA and RNA polymerase and other regulatory proteins. In this work, I review the process of transcription and selected mechanisms of its regulation, focusing on the effects of DNA topology. A key characteristic of DNA topology is the level of supercoiling. I describe how the DNA supercoiling influences initiation of transcription, and how it is affected by topoisomerases and nucleoid-associating proteins. I then discuss selected examples of regulation of gene expression by topology. Finally, I discuss the alternative σN factor from Bacillus subtilis, which allows more efficient transcription initiation from linear (i.e. relaxed) rather than supercoiled DNA templates. In this property, σN diametrically differs from other σ factors. Key words: DNA, topology, RNA polymerase, transcription, promoter, sigma factor
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Regulation of transcription in Streptococcus pneumoniae
Zajíčková, Adéla ; Doubravová, Linda (advisor) ; Hnilicová, Jarmila (referee)
Regulation of transcription in Streptococcus pneumoniae Streptococcus pneumoniae, an opportunistic human pathogen, encodes a single eukaryotic-like Ser/Thr kinase, StkP, which plays a crucial role in cell growth and division. In a phosphoproteomic study, StkP-dependent phosphorylation of SigAunder antibiotic- induced stress conditions was demonstrated in vivo. SigA, a housekeeping factor, in complex with RNA polymerase, is responsible for the transcription of most genes in S. pneumoniae. Therefore, phosphorylation of this important regulator might play an important role in transcriptional regulation. In this work, we confirmed that StkP phosphorylates SigA at residues T291 and T306 in vitro. However, we could not detect SigA phosphorylation in vivo, and the role of phosphorylation remains to be elucidated. We detected all RNAP subunits, including SigA, in the RNAP complex of S. pneumoniae by mass spectrometry analysis. The protein Spr0726 was identified as part of the RNAP complex. The reciprocal interaction between Spr0726 and RNAP was confirmed by immunoprecipitation experiments. In functional studies, we found that deletion of the spr0726 gene leads to a decrease in the amount of the subunits β and α in the RNAP complex. Our data suggest that the Spr0726 protein may affect the stability of the RNAP...
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Fourier transformation for exon prediction
Rusina, Michal ; Škutková, Helena (referee) ; Maděránková, Denisa (advisor)
In this bachelor thesis there are described the methods of the prediction of exon. The first part is aimed at the difference between the prokaryotic and eukaryotic organisms, the description of the DNA structure and the explanation of the terms exon and intron. The second section of the theoretic part includes four methods of the prediction of exons namely dynamic programming, neural networks, hidden Markov models and discrete Fourier transform. In the practical part there was created the program called Predikce_exonu that searches exons in nucleotide sequences and works on the principle of the Fourier transform. This algorithm together with 3 freely available programs was tested on 25 sequences and the success of their prediction was described by sensitivity and specificity.
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Web Application for Bioinformatics Education - Genetic Code
Kilián, Martin ; Martínek, Tomáš (referee) ; Burgetová, Ivana (advisor)
This thesis briefly describes basic informations in moleculary biology. Thesis deals with a transfer of information from DNA into RNA through uprotein, are also briefly described the methods and algorithms dealing with the detection of genome. The larger part deals with the recognition cue coded signals and determination of sequences and their subsequent transcription and translation, which demonstration program was needed to create. The final result is s web application as the education system.
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Gene Searching - Web Application
Stiborová, Lucie ; Koutný, Jiří (referee) ; Burgetová, Ivana (advisor)
The aim of my Bachelor's Thesis is to create a user interface and to implement the web application with the genes' searching. This is the educational application when each user has a possibility of entering own nucleotide sequences or selecting a redefined chain. This application has to be able to read, process and decide on the existence of a potential gene. On the base of this simple statistic method the result is given back to the user.
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Transcription motif finding algorithms
Peřinová, Barbora ; Škutková, Helena (referee) ; Maděránková, Denisa (advisor)
This bachelor thesis deals with finding transcription motifs in DNA sequences. The first part describes the DNA transcription process, the significance of transcription motifs and provides an overview of transcription motif databases. The second part contains the classification of transcription motif finding algorithms, and details seven existing algorithms. In the practical part, functions for the analysis according to the Oligo-Analysis algorithm were created. Created functions were validated on the artificially created sequences with introduced motifs. The analysis of two families of coregulated genes was realized and the results were compared with the values achieved by the authors of the Oligo-Analysis algorithm.
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Nucleotide density for finding CpG islands
Sikorová, Eva ; Provazník, Ivo (referee) ; Maděránková, Denisa (advisor)
This bachelor thesis deals with searching for CpG islands in the DNA sequences using the nucleotide density vectors. The first part includes view of the DNA structure, the genetic information expression, more detailed analysis of CpG islands and primarily their detection methods. In the practical part the algorithm for graphical representation of nucleotides on the basis of their densities and detections of CpG islands of artificially created and real DNA sequences was realized in the MATLAB program. The thesis includes the analysis of twenty sequences with the expected content of CpGIs and the comparison of results between the created program and two search engines.
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Tolerance of DNA damage by novel biologically active platinum complexes
Vystrčilová, Jana ; Vrána, Oldřich (referee) ; Nováková,, Olga (advisor)
The anti-tumor activity of platinum based drugs is mediated by their ability to attack DNA. Platinum complexes can alter the structure of DNA by modifying the bases, mainly guanines. The biological consequnces of such interactions are compromising replication and transcription. RNA polymerase complex can stall at a damaged site in DNA and mark the lesion for repair by proteins that are utilized to execute nucleotide excision repair, a pathway commonly associated with the removal of bulky DNA damage from the genome. This RNA polymerase-induced repair pathway is called transcription-coupled nucleotide excision repair. Main goal of this thesis was to study RNA polymerases tolerance of DNA damage by novel, biologically active platinum (II) complexes involving derivatives of aromatic cytokinines as the ligands; cis-[Pt(2-chloro-6-(4-methoxybenzylamino)-9-isopropylpurin)2Cl2](PR-001), cis-[Pt(2-chloro-6-(benzylamino)-9-isopropylpurin)2Cl2](PR-002 )and cis-[Pt(2-(3-hydroxypropylamino)-6-(benzylamino)-9-isopropylpurin)2Cl2](PR-005). DNA templates (constructs) that contain a single, site-specific DNA lesion and support transcription by human RNA polymerase II and bacteriophage T7 RNA polymerase were prepared. The method is making use of polymerase chain reaction (PCR) and biotin-streptavidin interactions and paramagnetic particles to purify the final product. Synthetic oligomers duplexes (75-mer, 56-mer and 15-mer) are ligated to 5´-biotin pCI-neo-G-lessT7 PCR fragment, the 15-mer is either unmodified or modified with a site-specific lesion of PR-005 and cisplatin. We also studied the inhibition of RNA polymerases activity on globally modified plasmid pCI-neo and pUC 19 by novel platinum complexes and cisplatin. We found that bifunctional adducts of complex PR-005 contrary to adducts of PR-001 and PR-002 effectively decrease amount of full lenght transcripts produced by both, human and bacterial RNA polymerases. This result can be explained by a sterical block, induced to DNA by intrastrand cross-link of PR-005 with bulky aromatic ligands.
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Phonetic transcription of Czech
Pavel, Lukáš ; Křupka, Aleš (referee) ; Sysel, Petr (advisor)
The goal is to create an application for automatic transcription of Czech language into a phonetic form. The first chapter of this thesis describes the basic phonetic units, alphabets and symbols, that phonetic alphabet uses. The second chapter describes in detail the rules of phonetic transcription of Czech language, including specific examples. Finally, in the third chapter is a description of the application created in C programming language, including flowcharts for better orientation in its functions.
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Role of the WIP1 phosphatase in the nucleolus
Palková, Natálie ; Macůrek, Libor (advisor) ; Sztacho, Martin (referee)
Protein phosphatase 2C delta (known as WIP1) is an important negative regulator of the DNA damage response (DDR) signalling. As a chromatin-bound protein, it dephosphorylates and thus inactivates ATM kinase and the transcription factor p53. Increased expression of WIP1 suppresses the function of the tumour suppressor p53 and contributes to the development of several cancer types, including breast and brain tumours. In recent years, it has been shown that WIP1 can also regulate cellular processes that are not directly linked to DDR, such as ensuring telomere stability. However, alternative functions of the WIP1 protein have not yet been fully understood. In this work, we described a novel role of the WIP1 phosphatase in the nucleolus, the organelle responsible for ribosome biogenesis. We found that WIP1 associates with many nuclear proteins, and using deletion mutants, we identified a nucleolar localisation sequence (NoLS) at the C-terminus of the protein. Using super-resolution microscopy, we detected the localisation of WIP1 phosphatase in the fibrillar centres of the nucleolus. We employed an inducible Cas9 system for generating double-strand breaks in ribosomal DNA and found that WIP1 has an impact on recruitment of DNA repair factors to the nucleolar caps. Analysis based on quantitative...
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