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Cryo-EM: Impact of EER format and super-resolution on SPA results
Geršl, David ; Dobrovský, Ladislav (referee) ; Hůlka, Tomáš (advisor)
Kryo-elektronová mikroskopie (cryo-EM) se stala zásadním nástrojem ve strukturální biologii, který umožňuje vizualizaci biologických makromolekul v atomovém rozlišení za kryogenních podmínek. Tato diplomová práce se věnuje zkoumání vlivu obrazového rozlišení a chyb lokalizace v rámci techniky cryo-EM, přičemž klade důraz na jejich dopad na analýzu jednotlivých částic (SPA). Práce je rozdělena do dvou hlavních experimentálních částí. První část zkoumá dopady různých obrazových rozlišení na procesy rekonstrukce v SPA. Zde bylo zjištěno, že zvýšení rozlišení obrazu vede k lepšímu detailu a strukturální jasnosti, avšak zároveň výrazně zvyšuje výpočetní nároky a komplexnost datového zpracování. Druhá experimentální část se zaměřuje na vliv uměle indukovaných chyb lokalizace, které jsou způsobeny použitím technik superrozlišení. Tyto chyby mají negativní vliv na přesnost korekce pohybu a odhady funkcí přenosu kontrastu (CTF), což jsou klíčové aspekty pro přesné 3D rekonstrukce v SPA. Zkoumání těchto dvou klíčových faktorů odhalilo, že i přes technologický pokrok v oblasti detektorů a zpracování obrazu, který umožnil dosáhnout rozlišení srovnatelného s X-ray krystalografií, stále existují výzvy spojené s potenciální optimalizací těchto procesů v praxi. Tyto výzvy zahrnují potřebu efektivnějšího využití výpočetních zdrojů a lepšího pochopení vlivu technických parametrů na kvalitu a přesnost výsledných rekonstrukcí. Výsledky této práce poskytují poznatky pro další vývoj v oblasti cryo-EM a otevírají možnosti pro zlepšení metod SPA, což může potenciálně vést k přesnějšímu a efektivnějšímu studiu komplexních biologických struktur. Tyto poznatky také naznačují směry pro budoucí výzkum a potenciál pro technologické inovace, které by mohly dále zlepšit schopnosti cryo-EM v oblasti strukturní biologie.
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Regularization methods for discrete inverse problems in single particle analysis
Havelková, Eva ; Hnětynková, Iveta (advisor) ; Plešinger, Martin (referee)
The aim of this thesis is to investigate applicability of regulariza- tion by Krylov subspace methods to discrete inverse problems arising in single particle analysis (SPA). We start with a smooth model formulation and describe its discretization, yielding an ill-posed inverse problem Ax ≈ b, where A is a lin- ear operator and b represents the measured noisy data. We provide theoretical background and overview of selected methods for the solution of general linear inverse problems. Then we focus on specific properties of inverse problems from SPA, and provide experimental analysis based on synthetically generated SPA datasets (experiments are performed in the Matlab enviroment). Turning to the solution of our inverse problem, we investigate in particular an approach based on iterative Hybrid LSQR with inner Tikhonov regularization. A reliable stopping criterion for the iterative part as well as parameter-choice method for the inner regularization are discussed. Providing a complete implementation of the proposed solver (in Matlab and in C++), its performance is evaluated on various SPA model datasets, considering high levels of noise and realistic distri- bution of orientations of scanning angles. Comparison to other regularization methods, including the ART method traditionally used in SPA,...
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Regularization methods for discrete inverse problems in single particle analysis
Havelková, Eva ; Hnětynková, Iveta (advisor)
The aim of this thesis is to investigate applicability of regulariza- tion by Krylov subspace methods to discrete inverse problems arising in single particle analysis (SPA). We start with a smooth model formulation and describe its discretization, yielding an ill-posed inverse problem Ax ≈ b, where A is a lin- ear operator and b represents the measured noisy data. We provide theoretical background and overview of selected methods for the solution of general linear inverse problems. Then we focus on specific properties of inverse problems from SPA, and provide experimental analysis based on synthetically generated SPA datasets (experiments are performed in the Matlab enviroment). Turning to the solution of our inverse problem, we investigate in particular an approach based on iterative Hybrid LSQR with inner Tikhonov regularization. A reliable stopping criterion for the iterative part as well as parameter-choice method for the inner regularization are discussed. Providing a complete implementation of the proposed solver (in Matlab and in C++), its performance is evaluated on various SPA model datasets, considering high levels of noise and realistic distri- bution of orientations of scanning angles. Comparison to other regularization methods, including the ART method traditionally used in SPA,...
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Regularization methods for discrete inverse problems in single particle analysis
Havelková, Eva ; Hnětynková, Iveta (advisor) ; Plešinger, Martin (referee)
The aim of this thesis is to investigate applicability of regulariza- tion by Krylov subspace methods to discrete inverse problems arising in single particle analysis (SPA). We start with a smooth model formulation and describe its discretization, yielding an ill-posed inverse problem Ax ≈ b, where A is a lin- ear operator and b represents the measured noisy data. We provide theoretical background and overview of selected methods for the solution of general linear inverse problems. Then we focus on specific properties of inverse problems from SPA, and provide experimental analysis based on synthetically generated SPA datasets (experiments are performed in the Matlab enviroment). Turning to the solution of our inverse problem, we investigate in particular an approach based on iterative Hybrid LSQR with inner Tikhonov regularization. A reliable stopping criterion for the iterative part as well as parameter-choice method for the inner regularization are discussed. Providing a complete implementation of the proposed solver (in Matlab and in C++), its performance is evaluated on various SPA model datasets, considering high levels of noise and realistic distri- bution of orientations of scanning angles. Comparison to other regularization methods, including the ART method traditionally used in SPA,...
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Evaluation of negative stains for single particle analysis in electron microscopy
ĎURINOVÁ, Eva
Four negative stains, hafnium chloride and europium, samarium and gadolinium nitrates, were tested in single particle electron microscopy as potential alternatives to uranyl acetate, which is recently being widely restricted for its toxicity. The new stains were applied to a structurally well-described plant photosystem I, visualized by a transmission electron microscope and classified in a single particle analysis. The quality of the stains was evaluated by the obtained resolution and ability to provide reliable structural information.
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