Download or read book Development of Site specific RNA Labeling Strategies to Probe Alternative RNA Splicing written by Tilemachos D. Kosmidis and published by . This book was released on 2016 with total page 588 pages. Available in PDF, EPUB and Kindle. Book excerpt: The objective of this project was to develop a method for the labeling of RNA at specific locations using non-natural base pairs. This thesis details the efforts made towards this objective through the development of a modular synthetic platform for an expanded genetic alphabet and its evaluation using transcriptional assays.Chapter 1 introduces the structure and function of RNA, followed by a description of RNA processing events with emphasis placed on alternative RNA splicing and how aberrations in splicing can lead to disease. The concept of RNA labeling follows and examples from the literature are presented along with challenges associated with these techniques to elucidate key mechanistic splicing concepts. The concept of orthogonal base pairs is then introduced along with representative examples from the literature. The application of this concept in RNA labeling is then presented along with key examples from the literature. The limitations of these methods are highlighted followed by the specific aims behind this project.Chapter 2 presents the efforts made towards the development of a modular synthetic route for the synthesis of C-ribonucleosides, with a Heck reaction as the key step. Specifically, its application in the synthesis of the Z ribonucleoside developed by the Benner group is investigated. The chapter starts with re-introducing the Z/P pair developed by the Benner group and highlighting the reasons that placed it as our chosen base pair for RNA labeling. The synthesis of C-deoxyribonucleosides utilizing the Heck reaction is presented, followed by the challenges associated with C-ribonucleoside synthesis.The syntheses of the alkene and halide coupling partners of the proposed Heck reaction are firstly presented.Investigations on the Heck reaction are then presented, followed by investigations on the transformation of nitro group in Z to moieties suitable for further derivatization. The key outcome of this work is that a synthetic route involving the Heck reaction for the synthesis of C-ribonucleosidesis not feasible. This is due to long reaction times required to drive the Heck reaction to completion and difficulties encountered with elucidation of its stereselectivity. Furthermore, PCR experiments conducted by our collaborators within the Eperon group at the University of Leicester revealed significant misincorporation of Z opposite G. Consequently, a change in strategy towards the use of nucleotides developed by the Hirao group was made.Chapter 3 describes the development of a robust synthetic route for the synthesis of a library of C6-functionalized 2-aminopurines as potential candidates for an expanded genetic alphabet in RNA. The s/Pa pair developed by the Hirao group is re-introduced.The installation of an alkyne functionality on the guanosine scaffold via a Sonogashira reaction is described,followed by investigations of [3+2] cycloaddition reactions between the alkyne and azides or aldehyde oximes. Finally, the development of a novel Suzuki-Miyaura protocol for the direct installation of heterocyclic substituents on the guanosine scaffold is also reported.Key outcomes of this chapter are the following. A robust method was developed for the expedient synthesis of C6-functionalized s analogues. This method enabled access to various classes of analogues in three or six steps,including triazoles, isoxazoles, thiophenes and pyrazoles. Attempts to install an azide moiety to the guanosine were partly successful, but the strategy was abanonded due to reaction reproducibility issues. In addition, a C-H activation strategy was not successful on installing an oxazole moiety Chapter 4 details the efforts towards the synthesis of nucleoside triphosphates based on the s analogues described in Chapter 3. This chapter will begin with the presentation of the most common strategies for the synthesis of nucleoside triphosphates. Initial attempts to synthesize triphosphates by global deprotection of nucleosides synthesized in Chapter 3 are next described. Attempts to install a silyl ether group in the 5’-OH are presented, followed by the installation of acetates on the 2’/3’ hydroxyls and attempts to protect the exocyclic amine on guanosine as the phenoxyacetamide. The installation of a pyrazole moiety via Suzuki-Miyauraprotocol using the corresponding boronic acid pinacol ester described in Chapter 3 is presented. The installation of an alkyne moiety is also described and the synthesis of an isoxazole analogue using this alkyne precursor will follow. The chapter will end with the presentation of the synthesis of pyrazole and isoxazole triphosphate analogues. The key outcome of this chapter is that employing different protecting groups on the 5’ and the 2’/3’ hydroxyls enabled the synthesis of pure triphosphates. Chapter 5 presents the evaluation of the nucleotide triphosphates synthesized in Chapter 4 regarding their transcriptional efficiency. Key outcome of this work are that high concentrations (>0.5 mM mM) are needed in order to observe significant incorporation of the analogues, compared to s, which needs 0.1 mM for efficient incorporation. At high concentration, the isoxazole moiety exhibits better incorporation efficiency compared to the pyrazole analogue. Furthermore, the addition of 0.5 mM of MnCl2 resulted in increased incorporation efficiency of the pyrazole analogue, while s and the isoxazole exhibited reduced efficiency under these conditions.
Download or read book Development of Site specific Labeling Strategies for Protein Modification and Synthesis of Rab Probes written by Long Yi and published by . This book was released on 2012 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Development of Site Specific Labeling Strategies for Protein Modification and Synthesis of Rab Probes written by Long Yi and published by . This book was released on 2012 with total page 181 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book RNA RNA Interactions written by Frank J. Schmidt and published by Humana Press. This book was released on 2014-10-29 with total page 219 pages. Available in PDF, EPUB and Kindle. Book excerpt: In this volume expert researchers in the field detail many of the methods which are now commonly used to study RNA. These methods are presented as a guidebook to scientists who are experienced with RNA research and want to brush up on a new technique. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and key tips on troubleshooting and avoiding known pitfalls. Thorough and intuitive, RNA-RNA Interactions: Methods and Protocols guides scientists investigating biological systems and studying RNA.
Download or read book Synthetic mRNA written by Robert E. Rhoads and published by Humana. This book was released on 2016-05-29 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: This volume presents detailed laboratory protocols for in vitro synthesis of mRNA with favorable properties, its introduction into cells by a variety of techniques, and the measurement of physiological and clinical consequences such as protein replacement and cancer immunotherapy. Synthetic techniques are described for structural features in mRNA that provide investigational tools such as fluorescence emission, click chemistry, photo-chemical crosslinking, and that produce mRNA with increased stability in the cell, increased translational efficiency, and reduced activation of the innate immune response. Protocols are described for clinical applications such as large-scale transfection of dendritic cells, production of GMP-grade mRNA, redirecting T cell specificity, and use of molecular adjuvants for RNA vaccines. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Synthetic mRNA: Production, Introduction into Cells, and Physiological Consequences is a valuable and cutting-edge resource for both laboratory investigators and clinicians interested in this powerful and rapidly evolving technology.
Download or read book RNA protein Interactions written by Kiyoshi Nagai and published by Oxford University Press, USA. This book was released on 1994 with total page 302 pages. Available in PDF, EPUB and Kindle. Book excerpt: The study of RNA-protein interactions is crucial to understanding the mechanisms and control of gene expression and protein synthesis. The realization that RNAs are often far more biologically active than was previously appreciated has stimulated a great deal of new research in this field. Uniquely, in this book, the world's leading researchers have collaborated to produce a comprehensive and current review of RNA-protein interactions for all scientists working in this area. Timely, comprehensive, and authoritative, this new Frontiers title will be invaluable for all researchers in molecular biology, biochemistry and structural biology.
Download or read book Modified Nucleic Acids written by Kazuhiko Nakatani and published by Springer. This book was released on 2016-04-04 with total page 280 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book spans diverse aspects of modified nucleic acids, from chemical synthesis and spectroscopy to in vivo applications, and highlights studies on chemical modifications of the backbone and nucleobases. Topics discussed include fluorescent pyrimidine and purine analogs, enzymatic approaches to the preparation of modified nucleic acids, emission and electron paramagnetic resonance (EPR) spectroscopy for studying nucleic acid structure and dynamics, non-covalent binding of low- and high-MW ligands to nucleic acids and the design of unnatural base pairs. This unique book addresses new developments and is designed for graduate level and professional research purposes.
Download or read book Ribozymes written by Sabine Müller and published by John Wiley & Sons. This book was released on 2021-07-09 with total page 81 pages. Available in PDF, EPUB and Kindle. Book excerpt: Ribozymes Provides comprehensive coverage of a core field in the molecular biosciences, bringing together decades of knowledge from the world’s top professionals in the field Timely and unique in its breadth of content, this all-encompassing and authoritative reference on ribozymes documents the great diversity of nucleic acid-based catalysis. It integrates the knowledge gained over the past 35 years in the field and features contributions from virtually every leading expert on the subject. Ribozymes is organized into six major parts. It starts by describing general principles and strategies of nucleic acid catalysis. It then introduces naturally occurring ribozymes and includes the search for new catalytic motifs or novel genomic locations of known motifs. Next, it covers the development and design of engineered ribozymes, before moving on to DNAzymes as a close relative of ribozymes. The next part examines the use of ribozymes for medicinal and environmental diagnostics, as well as for therapeutic tools. It finishes with a look at the tools and methods in ribozyme research, including the techniques and assays for structural and functional characterization of nucleic acid catalysts. The first reference to tie together all aspects of the multi-faceted field of ribozymes Features more than 30 comprehensive chapters in two volumes Covers the chemical principles of RNA catalysis; naturally occurring ribozymes, engineered ribozymes; DNAzymes; ribozymes as tools in diagnostics and therapy, and tools and methods to study ribozymes Includes first-hand accounts of concepts, techniques, and applications by a team of top international experts from leading academic institutions Dedicates half of its content to methods and practical applications, ranging from bioanalytical tools to medical diagnostics to therapeutics Ribozymes is an unmatched resource for all biochemists, biotechnologists, molecular biologists, and bioengineers interested in the topic.
Download or read book Transcription and Splicing written by B. D. Hames and published by Oxford University Press, USA. This book was released on 1988 with total page 238 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book gives a co-ordinated review of our present knowledge of eukaryotic RNA synthesis.
Download or read book Probes and Tags to Study Biomolecular Function written by Lawrence W. Miller and published by John Wiley & Sons. This book was released on 2008-09-08 with total page 193 pages. Available in PDF, EPUB and Kindle. Book excerpt: The only resource to systematically review current experimental methods, this handy reference enables researchers to select the best solution for their experimental problems. For each method covered, the book provides step-by-step protocols, illustrated by typical research applications. After an initial section on probing the lipid bilayer, the text moves on to discuss probing proteins -- including membrane proteins -- and nucleic acids. The first single publication to incorporate chemical markers, fluorescent probes and genetic tags allows a well-informed comparison of different solutions for the same problem in molecular analytics.
Download or read book Molecular Biology of The Cell written by Bruce Alberts and published by . This book was released on 2002 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book Plant RNA Biology written by Dóra Szakonyi and published by Frontiers Media SA. This book was released on 2019-11-18 with total page 167 pages. Available in PDF, EPUB and Kindle. Book excerpt: Discoveries from the past decades revealed that RNA molecules are much more than inert intermediates between the coding DNA sequences and their functional products, proteins. Today, RNAs are recognized as active regulatory molecules influencing gene expression, chromatin organization and genome stability, thus impacting all aspects of plant life including development, growth, reproduction and stress tolerance. Innovations in methodologies, the expanding application of next-generation sequencing technologies, and the creation of public datasets and databases have exposed a new universe of RNA-based mechanisms and led to the discovery of new families of non-coding RNAs, uncovered the large extent of alternative splicing events, and highlighted the potential roles of RNA modifications and RNA secondary structures. Furthermore, considerable advances have been made in identifying RNA-binding and processing factors involved in the synthesis and maturation of different forms of RNA molecules as well as in RNA processing, biochemical modifications or degradation. This Research Topic showcases the broad biological significance of RNAs in plant systems and contains eight original research articles, one review and four mini-reviews, covering various RNA-based mechanisms in higher plants. Emerging new technologies and novel multidisciplinary approaches are empowering the scientific community and will expectedly bring novel insights into our understanding of the mechanisms through which RNA is regulated and regulates biological processes in plant cells.
Download or read book New Strategies in Chemical Synthesis and Catalysis written by Bruno Pignataro and published by John Wiley & Sons. This book was released on 2012-09-24 with total page 407 pages. Available in PDF, EPUB and Kindle. Book excerpt: This volume represents one of the two edited by inviting a selection of young researchers participating to the European Young Chemist Award 2010. The other volume concerns the area of Nanotechnology/Material Science and is titled: Molecules at Work. This book contains the contributions of selected young chemists from the field of synthetic chemistry. The contributions are grouped under the three following umbrella topics: Synthetic Methods Catalysis Combinatorial and Chemical Biology This volume is an indispensable read for all organic and inorganic chemists, biochemists, chemists working with/on organometallics, and Ph.D. students in chemistry interested in seeing what tomorrow's chemistry will look like.
Download or read book Small Molecule Microarrays Methods and Protocols written by Mahesh Uttamchandani and published by Methods in Molecular Biology. This book was released on 2018-12-05 with total page 274 pages. Available in PDF, EPUB and Kindle. Book excerpt:
Download or read book RNA Tagging written by Manfred Heinlein and published by Humana. This book was released on 2021-08-08 with total page 490 pages. Available in PDF, EPUB and Kindle. Book excerpt: This book provides a compendium of state-of-the-art methods for the labeling, detection, and purification of RNA and RNA-protein complexes and thereby constitutes an important toolbox for researchers interested in understanding the complex roles of RNA molecules in development, signaling, and disease. Beginning with a section on in situ detection of RNA molecules using FISH techniques, the volume continues with parts exploring in vivo imaging of RNA transport and localization, imaging and analysis of RNA uptake and transport between cells, identification and analysis of RNA-binding proteins, guide RNAs in genome editing, as well as other specific analytical techniques. Written for the highly successful Methods in Molecular Biology series, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols, and tips on troubleshooting and avoiding known pitfalls. Authoritative and practical, RNA Tagging: Methods and Protocols serves as a vital reference for researchers looking to further the increasingly important research in RNA biology.
Download or read book DNA and RNA Modification Enzymes written by Henri Grosjean and published by CRC Press. This book was released on 2009-06-11 with total page 682 pages. Available in PDF, EPUB and Kindle. Book excerpt: This volume is a timely and comprehensive description of the many facets of DNA and RNA modification-editing processes and to some extent repair mechanisms. Each chapter offers fundamental principles as well as up to date information on recent advances in the field (up to end 2008). They ended by a shortconclusion and future prospect' section and
Download or read book RNA written by Donald Charles Rio and published by . This book was released on 2011 with total page 0 pages. Available in PDF, EPUB and Kindle. Book excerpt: So much has been learned about RNA in the past ten years that the ability to purify, analyze, and manipulate RNA molecules is now essential in all kinds of bioscience. Initiating RNA research can be intimidating but the new book RNA: A Laboratory Manualprovides a broad range of up-to-date techniques presented in a functional framework, so that any investigator can confidently handle RNA and carry out meaningful experiments, from the most basic to the highly sophisticated. Originating in three of the field's most prominent laboratories, this manual provides the necessary background and strategies for approaching any RNA investigation, as well as detailed protocols and extensive tips and troubleshooting information. It is required reading for every research laboratory in the life sciences.
