| Edizione | [Second edition.] |
| Pubbl/distr/stampa |
Weinheim : , : John Wiley & Sons, Inc., , 2014
|
| Descrizione fisica |
1 online resource (xlii, 1314 pages)
|
| Disciplina |
574.192
|
| Soggetto topico |
Biochemistry
Biochemistry - Outlines, syllabi, etc
|
| ISBN |
3-527-64706-6
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| Formato |
Materiale a stampa  |
| Livello bibliografico |
Monografia |
| Lingua di pubblicazione |
eng
|
| Nota di contenuto |
Cover; Title Page; Copyright; Contents; Preface; List of Contributors; Part I RNA Synthesis and Detection; Chapter 1 Enzymatic RNA Synthesis Using Bacteriophage T7 RNA Polymerase; 1.1 Introduction; 1.2 Description of Method -- T7 Transcription In vitro; 1.2.1 Templates; 1.2.1.1 Strategy (i): Insertion into a Plasmid; 1.2.1.2 Strategy (ii): Direct Use of Templates Generated by PCR; 1.2.1.3 Strategy (iii): Annealing of a T7 Promoter DNA Oligonucleotide to a Single-Stranded Template; 1.2.2 Special Demands on the RNA Product 1.2.2.1 Homogeneous 5' and 3' Ends, Small RNAs, Functional Groups at the 5' End1.2.2.2 Modified Substrates; 1.3 Transcription Protocols; 1.3.1 Transcription with Unmodified Nucleotides; 1.3.2 Transcription with 2' -Fluoro-Modified Nucleotides; 1.3.3 T7 Transcripts with 5' -- Cap Structures; 1.3.4 Purification; 1.4 Troubleshooting; 1.4.1 Low or No Product Yield; 1.5 Rapid Preparation of T7 RNA Polymerase; 1.5.1 Required Material; 1.5.1.1 Medium; 1.5.1.2 Buffers and Solutions; 1.5.1.3 Electrophoresis and Chromatography; 1.5.2 Procedure 1.5.2.1 Cell Growth, Induction, and Test for Expression of T7 RNAP1.5.2.2 Purification of T7 RNAP; 1.5.3 Notes and Troubleshooting; References; Chapter 2 Production of RNAs with Homogeneous 5' -- and 3' -Ends; 2.1 Introduction; 2.2 Description of Approach; 2.2.1 Cis-Cleaving Autocatalytic Ribozyme Cassettes; 2.2.1.1 The 5' -Cassette; 2.2.1.2 The 3' -Cassette; 2.2.1.3 Purification of Released RNA Product and Conversion of End Groups; 2.2.2 Trans-Cleaving Ribozymes for the Generation of Homogeneous 3' Ends; 2.2.3 Further Strategies toward Homogeneous Ends 2.3 Critical Experimental Steps, Changeable Parameters, Troubleshooting2.3.1 Construction of Cis-Cleaving 5'- and 3'-Cassettes; 2.4 PCR Protocols; 2.5 Potential Problems; References; Chapter 3 RNA Ligation; 3.1 General Introduction; 3.1.1 T4 Polynucleotide Ligases; 3.1.2 Reaction Mechanism; 3.1.3 Advantages of T4 DNA Ligase for RNA Ligation; 3.1.4 Chapter Structure; 3.2 RNA Ligation Using T4 DNA Ligase (T4 Dnl); 3.2.1 Overview of the RNA Ligation Method Using the T4 DNA Ligase (T4 Dnl); 3.2.2 Large-Scale Transcription and Purification of RNAs 3.2.3 Generating Homogeneous Acceptor 3' -Ends for Ligation3.2.4 Site-Directed Cleavage with RNase H; 3.2.5 Dephosphorylation and Phosphorylation of RNAs; 3.2.6 RNA Ligation; 3.2.7 Troubleshooting; 3.3 Simultaneous Splint Ligation of Five RNA Fragments to Generate RNAs for FRET Experiments; 3.3.1 Introduction; 3.3.2 Construct Design; 3.3.3 Troubleshooting; 3.3.3.1 Low Overall Ligation Efficiency; 3.3.3.2 Undesired Ligation By-products; 3.3.3.3 RNA Degradation; 3.4 T4 RNA Ligase(s); 3.4.1 Introduction; 3.4.2 Mechanism and Substrate Specificity; 3.4.2.1 Early Studies.
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| Altri titoli varianti |
Handbook of RNA Biochemistry: Second, Completely Revised and Enlarged Edition
Handbook of RNA Biochemistry
|
| Record Nr. | UNINA-9910830656003321 |
Info
Handbook of RNA biochemistry / / edited by Roland K. Hartmann