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Genetic interactions of the Prdm9 gene
Šebestová, Lenka ; Trachtulec, Zdeněk (advisor) ; Král, Jiří (referee)
The Prdm9 gene (PR domain containing 9, Meisetz, Hybrid sterility 1) encodes enzyme that trimethylates histone 3 on lysines 4 and 36. These methylation marks determine the positions of DNA double-strand breaks that are repaired by meiotic homologous recombination. In this study, we assayed genetic interactions of Prdm9 with two genes important for spermatogenesis - Mili (Piwil2) involved in piRNA biogenesis and Mybl1 encoding transcription factor that regulates many genes important for prophase I, including piRNA precursors. We crossed laboratory mice carrying mutation in Prdm9 with heterozygotes for mutation in Mybl1 or Mili, and created compound heterozygotes and, in case of Mybl1, also double homozygotes. We assessed body weight and male fertility parameters (weight of testes, sperm count, malformed sperm, percentage of tubules containing spermatocytes and of abnormal nuclei of pachytene spermatocytes) of these mice and compared them to controls. We also investigated the effect of Mybl1 and Mili mutations on fecundity of F1 intersubspecific hybrids. Our results revealed possible interactions of Prdm9 and Mybl1 in the laboratory mouse. Decreased gene dosage of Mybl1 reduced fertility of intersubspecific F1 hybrids. Interaction between Prdm9 and Mili in both laboratory mouse and F1 hybrids remain...
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The role of cohesin genes in the meiosis of male house mouse
Šebestová, Lenka ; Trachtulec, Zdeněk (advisor) ; Král, Jiří (referee)
Cohesin genes play an important role in cell division. They ensure proper chromosome segregation during mitosis and meiosis. This study is focused on the role of cohesin genes during meiosis in male house mouse (Mus musculus). At first, this study introduces key processes of mammalian meiosis. Next, the structure of cohesin complex is described; it consists of a heterodimer SMC proteins - SMC3 and SMC1α or SMC1β, which are enclosed to the ring by cleavable subunit RAD21, RAD21L or REC8. Fourth subunit - a STAG protein (STAG1, STAG2 or STAG3) associates with the cleavable subunit. Meiotic function of specific cohesin proteins (SMC1β, RAD21L, REC8 and STAG3) as deduced from the phenotypes of the deficiencies of their genes in male mouse is depicted. All these four genes are necessary for many processes during meiosis, - e.q. sister chromatid cohesion maintenance, synapsis and recombination. STAG3, SMC1β, and REC8 are necessary for centromeric cohesion. STAG3 and RAD21L are important for the assembly of the remaining cohesin subunits. The most important phenotype of deficiency of all four genes is the complete meiotic arrest in male prophase I. Therefore, cohesin research is important for the investigation of the causes of sterility in mammals. key words: cohesin, meiosis, spermatogenesis, mouse,...
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Biologically Active Metabolites of Plants. III. Alkaloids from Chelidonium Majus L. and Their Neurotropic and Antioxidant Activity
Šebestová, Lenka ; Cahlíková, Lucie (advisor) ; Opletal, Lubomír (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Departement: Department of Pharmaceutical Botany and Ecology Candidate: Mgr. Lenka Šebestová Adviser: Ing. Lucie Cahlíková, PhD. Thesis title: Biologically active metabolites of plants. III. Alkaloids from Chelidonium majus L. and their neurotropic and antioxidant activity. The aim of this thesis, which followed the diploma thesis, was final treatment of alkaloid fraction of benzophenanthridine alkaloids from Chelidonium majus, the isolation of minimum one alkaloid in pure form and to determine of its cholinesterase and antioxidative activity. Using column chromatography, prep. TLC and crystallization were isolated two benzophanathridine alkaloids marked as LŠ3 and LŠ4. First compound LŠ3 has been identified by comparation of their spectral data with those reported in the literature. LŠ4 has been identified on the basis of thein MS, NMR spectra and consultations. The activity of both alkaloids again erythrocyte acetylcholinesterase (HuAChE) and plasma butyrylcholinesterase (HuBuChE) were determined. Values of IC50 were calculated for both alkaloids (IC50 (HuAChE) = 3,25 ± 0,24 µM, IC50 (HuBuChE) = 4,51 ± 0,31 µM, 6-ethoxydihydrochelerythrin: IC50 (HuAChE) = 0,83 ± 0,04 µM, IC50 Hu(BuChE) = 4,20 ± 0,19 µM). Both alkaloids showed...
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Genetic interactions of the Prdm9 gene
Šebestová, Lenka ; Trachtulec, Zdeněk (advisor) ; Král, Jiří (referee)
The Prdm9 gene (PR domain containing 9, Meisetz, Hybrid sterility 1) encodes enzyme that trimethylates histone 3 on lysines 4 and 36. These methylation marks determine the positions of DNA double-strand breaks that are repaired by meiotic homologous recombination. In this study, we assayed genetic interactions of Prdm9 with two genes important for spermatogenesis - Mili (Piwil2) involved in piRNA biogenesis and Mybl1 encoding transcription factor that regulates many genes important for prophase I, including piRNA precursors. We crossed laboratory mice carrying mutation in Prdm9 with heterozygotes for mutation in Mybl1 or Mili, and created compound heterozygotes and, in case of Mybl1, also double homozygotes. We assessed body weight and male fertility parameters (weight of testes, sperm count, malformed sperm, percentage of tubules containing spermatocytes and of abnormal nuclei of pachytene spermatocytes) of these mice and compared them to controls. We also investigated the effect of Mybl1 and Mili mutations on fecundity of F1 intersubspecific hybrids. Our results revealed possible interactions of Prdm9 and Mybl1 in the laboratory mouse. Decreased gene dosage of Mybl1 reduced fertility of intersubspecific F1 hybrids. Interaction between Prdm9 and Mili in both laboratory mouse and F1 hybrids remain...
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The role of cohesin genes in the meiosis of male house mouse
Šebestová, Lenka ; Trachtulec, Zdeněk (advisor) ; Král, Jiří (referee)
Cohesin genes play an important role in cell division. They ensure proper chromosome segregation during mitosis and meiosis. This study is focused on the role of cohesin genes during meiosis in male house mouse (Mus musculus). At first, this study introduces key processes of mammalian meiosis. Next, the structure of cohesin complex is described; it consists of a heterodimer SMC proteins - SMC3 and SMC1α or SMC1β, which are enclosed to the ring by cleavable subunit RAD21, RAD21L or REC8. Fourth subunit - a STAG protein (STAG1, STAG2 or STAG3) associates with the cleavable subunit. Meiotic function of specific cohesin proteins (SMC1β, RAD21L, REC8 and STAG3) as deduced from the phenotypes of the deficiencies of their genes in male mouse is depicted. All these four genes are necessary for many processes during meiosis, - e.q. sister chromatid cohesion maintenance, synapsis and recombination. STAG3, SMC1β, and REC8 are necessary for centromeric cohesion. STAG3 and RAD21L are important for the assembly of the remaining cohesin subunits. The most important phenotype of deficiency of all four genes is the complete meiotic arrest in male prophase I. Therefore, cohesin research is important for the investigation of the causes of sterility in mammals. key words: cohesin, meiosis, spermatogenesis, mouse,...
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Biologically Active Metabolites of Plants. III. Alkaloids from Chelidonium Majus L. and Their Neurotropic and Antioxidant Activity
Šebestová, Lenka ; Cahlíková, Lucie (advisor) ; Opletal, Lubomír (referee)
Charles University in Prague, Faculty of Pharmacy in Hradec Králové Departement: Department of Pharmaceutical Botany and Ecology Candidate: Mgr. Lenka Šebestová Adviser: Ing. Lucie Cahlíková, PhD. Thesis title: Biologically active metabolites of plants. III. Alkaloids from Chelidonium majus L. and their neurotropic and antioxidant activity. The aim of this thesis, which followed the diploma thesis, was final treatment of alkaloid fraction of benzophenanthridine alkaloids from Chelidonium majus, the isolation of minimum one alkaloid in pure form and to determine of its cholinesterase and antioxidative activity. Using column chromatography, prep. TLC and crystallization were isolated two benzophanathridine alkaloids marked as LŠ3 and LŠ4. First compound LŠ3 has been identified by comparation of their spectral data with those reported in the literature. LŠ4 has been identified on the basis of thein MS, NMR spectra and consultations. The activity of both alkaloids again erythrocyte acetylcholinesterase (HuAChE) and plasma butyrylcholinesterase (HuBuChE) were determined. Values of IC50 were calculated for both alkaloids (IC50 (HuAChE) = 3,25 ± 0,24 µM, IC50 (HuBuChE) = 4,51 ± 0,31 µM, 6-ethoxydihydrochelerythrin: IC50 (HuAChE) = 0,83 ± 0,04 µM, IC50 Hu(BuChE) = 4,20 ± 0,19 µM). Both alkaloids showed...
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Biologically active metabolites of plants. 3. Alkaloids from Chelidonium majus L. and their inhibiting activity to acetycholinesterase.
Šebestová, Lenka ; Siatka, Tomáš (referee) ; Cahlíková, Lucie (advisor)
Šebestová L.: Biologically active metabolites of plants. 3. Alkaloids from Chelidonium majus L. and their inhibiting activity to acetylcholinesterase. Diploma thesis, Charles University in Prague, Faculty of Pharmacy in Hradec Králové, Department of Pharmaceutical Botany and Ecology, Hradec Králové 2010, 68 p. The thesis aims to process the shake-out of benzophenanthridine alkaloids, of Chelidonium majus, to isolate at least one alkaloid in pure form; consequently the thesis aims to give exact figures for the anti-cholinesterase, anti-protozoal a antioxidant activity of the alkaloid. The method of column chromatography was used to isolate the two alkaloids of the shake-out. The isolated substances were labelled LŠ1 and LŠ2 respectively. The identification of the substances as stylopine.and chelidonine respectively, was based on NMR and MS studies. Both isolated alkaloids were subjected to inhibition study with respect to human erytrocytal acetylcholinesterase and human seral butyrylcholinesterase. Stylopin showed the following set of results: IC50 (AChE) = 114 ± 2.9 µM, IC50 (BuChE) = >1000 µM. The results being considerably higher in comparison with the standard used, no further usage for the treatment of Alzheimer's disease is expected. Chelidonine showed the following set of results: IC50 (AChE)...
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