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The discovery of catalytic RNAs in the mid-1980s marked the beginning of a new era in RNA biology and an ever increasing appreciation of the diverse and critical roles played by this fascinating molecule. In Recombinant and In Vitro RNA Synthesis: Methods and Protocols, expert researchers in the field detail approaches from the inception of a new RNA project to the final sample ready for analysis. These experimental protocols combine established work-horse approaches with some of the latest innovative methods for RNA synthesis and purification. Written in the highly successful Methods in Molecular Biology(TM) 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.
Authoritative and practical, Recombinant and In Vitro RNA Synthesis: Methods and Protocols aids scientists in continuing to study the fascinating world of RNA.
List of contents
Purification of RNA Expressed in vivo Inserted in a tRNA Scaffold.-Selective RNase H Cleavage of Target RNAs from a tRNA Scaffold.- Preparation of Long Templates for RNA in vitro Transcription by Recursive PCR.- General Protocols for Preparation of Plasmid DNA Template, RNA in vitro Transcription and RNA Purification by Denaturing PAGE.- Preparation of Short RNA by in vitro Transcription.- Native RNA Purification by Gel Filtration Chromatography.- Cis-acting Ribozymes for the Production of RNA in vitro Transcripts with Defined 5' and 3' Ends.- Trans-acting Antigenomic HDV Ribozyme for Production of in vitro Transcripts with Homogenous 3' Ends.- Rapid Preparation of RNA Samples using DNA-affinity Chromatography and DNAzyme Methods.-Preparation of lN-GST Fusion Protein for Affinity Immobilization of RNA.- Affinity Purification of RNA using an ARiBo Tag.- Plasmid Template Design and in vitro Transcription of Short RNAs within a 'Structure Cassette' for Structure Probing Experiments.- In Vitro Transcription of Modified RNAs.- End-Labeling Oligonucleotides with Chemical Tags After Synthesis.- High-Purity Enzymatic Synthesis of Site-Specifically Modified tRNA.- Se-derivatized RNAs for X-ray Crystallography.- Biosynthetic Preparation of 13C/15N-labelled rNTPs for High-resolution NMR Studies of RNAs.- Preparative Separation of Ribonucleoside Monophosphates by Ion-Pair Reverse-Phase HPLC.- Splint Ligation of RNA with T4 DNA Ligase.
Summary
The discovery of catalytic RNAs in the mid-1980s marked the beginning of a new era in RNA biology and an ever increasing appreciation of the diverse and critical roles played by this fascinating molecule. In Recombinant and In Vitro RNA Synthesis: Methods and Protocols, expert researchers in the field detail approaches from the inception of a new RNA project to the final sample ready for analysis. These experimental protocols combine established work-horse approaches with some of the latest innovative methods for RNA synthesis and purification. 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.
Authoritative and practical, Recombinant and In Vitro RNA Synthesis: Methods and Protocols aids scientists in continuing to study the fascinating world of RNA.
