Oligonucleotide Synthesis: Methods and Applications

ISBN-13: 9781617374418 / Angielski / Miękka / 2010 / 436 str.

Piet Herdewijn

Opis

A collection of powerful new techniques for oligonucleotide synthesis and for the use of modified oligonucleotides in biotechnology. Among the protocol highlights are a novel two-step process that yields a high purity, less costly, DNA, the synthesis of phosphorothioates using new sulfur transfer agents, the synthesis of LNA, peptide conjugation methods to improve cellular delivery and cell-specific targeting, and triple helix formation. The applications include using molecular beacons to monitor the PCR amplification process, nuclease footprinting to study the sequence-selective binding of small molecules of DNA, nucleic acid libraries, and the use of small interference RNA (siRNA) as an inhibitor of gene expression.

Spis treści

Synthesis of DNA Using a New Two-Step Cycle Douglas J. Dellinger, Jason R. Betley, Tadeusz K. Wyrzykiewicz, and Marvin H. Caruthers RNA Synthesis Using 2'-O-(Tert-Butyldimethylsilyl) Protection Brian S. Sproat RNA Oligonucleotide Synthesis Via 5'-Silyl-2'-Orthoester Chemistry Stephanie A. Hartsel, David E. Kitchen, Stephen A. Scaringe, and William S. Marshall Dimethylthiarum Disulfide: New Sulfur Transfer Reagent in Phosphorothioates Oligonucleotide Synthesis Zhiwei Wang, Quanlai Song, and Yogesh S. Sanghvi Assay for Evaluating Ribonuclease H-Mediated Degradation of RNA-Antisense Oligonucleotide Duplexes Annie Galarneau, Kyung-Lyum Min, Maria M. Mangos, and Masad J. Damha Di- and Oligonucleotide Synthesis Using H-Phosphonate Chemistry Jacek Stawinski and Roger Strömberg Solid-Phase Synthesis of Circular Oligonucleotides Enrique Pedroso, Nuria Escaja, Miriam Frieden, and Anna Grandas Locked Nucleic Acid Synthesis Henrik M. Pfundheller, Anders M. Sørensen, Christian Lomholt, Anders M. Johansen, Troels Koch, and Jesper Wengel Synthesis of DNA Mimics Representing HypNA-pPNA Hetero-oligomers Vladimir A. Efimov and Oksana G. Chakhmakhcheva Base-Modified Oligonucleotides With Increased Duplex Stability: Pyrazolo[3,4-d]pyrimidines Replacing Purines Frank Seela, Yang He, Junlin He, Georg Becher, Rita Kröschel, Matthias Zulauf, and Peter Leonard Introduction of Hypermodified Nucleotides in RNA Darrell R. Davis and Ashok C. Bajji Chemical Methods for Peptide-Oligonucleotide Conjugate Synthesis Dmitry A. Stetsenko and Michael J. Gait Biotin-Labeled Oligonucleotides With Extraordinarily Long Tethering Arms A. Michael Morocho, Valeri Karamyshev, Olga Shcherbinina, and Nikolai Polushin Universal Labeling Chemistry for Nucleic Acid Detection on DNA Chips Ali Laayoun,Eloy Bernal-Méndez, Isabelle Sothier, Mitsuharu Kotera, and Alain Troesch Postsynthetic Functionalization of Triple Helix-Forming Oligonucleotides Alexandre S. Boutorine, and Jian-Sheng Sun Fluorescence-Based On-Line Detection as an Analytical Tool in RNA Electrophoresis Ute Scheffer and Michael Göbel Design and Optimization of Molecular Beacon Real-Time Polymerase Chain Reaction Assays Jacqueline A. M. Vet and Salvatore A. E. Marras High-Throughput Production of Optimized Primers (Fimers) for Whole-Genome Direct Sequencing Nikolai Polushin, Andrei Malykh, A. Michael Morocho, Alexei Slesarev, and Sergei Kozyavkin Nonenzymatic Template-Directed RNA Synthesis Marcus Hey and Michael Göbel DNAse I Footprinting of Small Molecule Binding Sites on DNA Christian Bailly, Jérôme Kluza, Christopher Martin, Thomas Ellis, and Michael J. Waring Gene Targeting Using Peptide Nucleic Acid Peter E. Nielsen Nucleic Acid Library Construction Using Synthetic DNA Constructs Hani S. Zaher and Peter J. Unrau In Vitro Selection From Combinatorial Nucleic Acid Libraries Andres Jäschke In Vitro Selection Procedures for Identifying DNA and RNA Aptamers Targeted to Nucleic Acids and Proteins Eric Dausse, Christian Cazenave, Bernard Rayner, and Jean-Jacques Toulmé Short Double-Stranded Ribonucleic Acid as Inhibitor of Gene Expression by the Interference Mechanism Jean-Rémi Bertrand, Andrei Maksimenko, and Claude Malvy Index

O autorze

Piet Herdewijn is Professor at the Katholieke Universiteit Leuven, Belgium, and head of the laboratory of medicinal chemistry at the Rega Institute. He obtained his Ph.D. and habilitation from the same university. He was a postdoctoral fellow of the Humboldt Foundation at the University of Konstanz. He has conducted research in bioorganic and medicinal chemistry, mainly in the field of nucleosides, nucleotides, and nucleic acids, at the University of Leuven, the University of Ghent, and the Université d'Evry-Val-d'Essonne.

Blurb

Nucleic acids chemistry has progressed rapidly in the last 10 years with the introduction of many new techniques and the development of many novel therapeutic entities. In Oligonucleotide Synthesis: Methods and Applications, laboratory experts describe in step-by-step detail the powerful new techniques they have developed for oligonucleotide synthesis and for the use of modified oligonucleotides in biotechnology. Among the protocol highlights are the synthesis of phosphorothioates via new sulfur transfer agents, the synthesis of LNA, peptide conjugation methods to improve cellular delivery and cell-specific targeting, triple helix formation, and a novel two-step process that yields a high purity DNA that is less costly. Novel applications include the use of molecular beacons to monitor the PCR amplification process, nuclease footprinting to study the sequence-selective binding of small molecules to DNA, nucleic acid libraries, optimization of RNA synthesis for siRNA applications, and the use of small interference RNA (siRNA) as an inhibitor of gene expression. The protocols follow the successful Methods in Molecular Biology™ series format, each one offering step-by-step laboratory instructions, an introduction outlining the principle behind the technique, lists of the necessary equipment and reagents, and tips on troubleshooting and avoiding known pitfalls.
Cutting-edge and highly practical, Oligonucleotide Synthesis: Methods and Applications offers today's investigators not only insight into important new biotechnologies, but also a full range of the detailed protocols needed to work successfully in DNA synthesis and its applications today.