Alternative pre-mRNA Splicing: Theory and Protocols - download pdf or read online

By Stefan Stamm, Chris Smith, Reinhard Lührmann

ISBN-10: 3527326065

ISBN-13: 9783527326068

ISBN-10: 3527636773

ISBN-13: 9783527636778

The publication contains significant components: The first one presents a quick theoretical
introduction that provides a quick review of different splicing and cites key papers within the field for extra in-depth info. the second one half is a set of experimental protocols which are utilized in the field of other splicing.
Contents
1 Splicing within the RNA World
2 RNPs, Small RNAs, and miRNAs
3 RNA components curious about Splicing
4 A Structural Biology viewpoint of Proteins eager about Splicing Regulation
5 The Spliceosome in Constitutive Splicing
6 using Saccharomyces cerevisiae to review the Mechanism of pre-mRNA Splicing
7 demanding situations in Plant substitute Splicing
8 substitute Splice web site Selection
9 Integration of Splicing with Nuclear and mobile Events
10 Splicing and Disease
11 From Bedside to Bench: the best way to learn a Splicing Mutation
12 research of universal Splicing Problems
13 Ultracentrifugation within the research and Purification of Spliceosomes Assembled In Vitro
14 Chemical Synthesis of RNA
15 RNA Interference (siRNA, shRNA)
16 Expression and Purification of Splicing Proteins
17 Detection of RNA–Protein Complexes via Electrophoretic Mobility Shift Assay
18 sensible research of enormous Exonic Sequences via Iterative In Vivo Selection
19 identity of Splicing cis-Elements via an Ultra-Refined Antisense Microwalk
20 Genomic SELEX to spot RNA pursuits of Plant RNA-Binding Proteins
21 Quantification of different Splice Variants
22 High-Throughput research of other Splicing through RT-PCR
23 tracking alterations in Plant replacement Splicing Events
24 Array Analysis
25 The CLIP technique to research Protein–RNA Interactions in Intact Cells and Tissues
26 RNA–Protein Crosslinking and Immunoprecipitation (CLIP) in Schizosaccharomyces pombe
27 identity of Proteins guaranteed to RNA
28 Single-Cell Detection of Splicing occasions with Fluorescent Splicing Reporters
29 The guidance of HeLa mobile Nuclear Extracts
30 In Vitro Splicing Assays
31 meeting and Isolation of Spliceosomal Complexes In Vitro
32 research of Site-Specific RNA–Protein Interactions
33 Immunoprecipitation and Pull-Down of Nuclear Proteins
34 research of Protein (-RNA) Complexes via (Quantitative) Mass Spectrometric Analysis
35 quick Cloning of Splicing Reporter Minigenes
36 In Vivo research of Splicing Assays
37 Coupled Promoter Splicing Systems
38 good telephone traces with Splicing Reporters
39 Splicing issue ChIP and ChRIP: Detection ofSplicing and Splicing components at
Genes by way of Chromatin Immunoprecipitation
40 Yeast Genetics to enquire the functionality of middle Pre-mRNA Splicing Factors
41 research of HIV-1 RNA Splicing
42 In Vivo research of Plant Intron Splicing
43 amendment State-Specific Antibodies
44 research of other Splicing in Drosophila Genetic Mosaics
45 Antisense Derivatives of U7 Small Nuclear RNA as Modulators of Pre-mRNA Splicing
46 Screening for substitute Splicing Modulators
47 Use of Oligonucleotides to alter Splicing
48 altering indications to the Spliceosome
49 assessment of Splicing proper Databases
50 research of RNA Transcripts by way of High-Throughput RNA Sequencing
51 id of Splicing issue goal Genes by means of High-Throughput Sequencing
52 Bioinformatic research of Splicing Events
53 research of Pre-mRNA Secondary constructions and substitute Splicing
54 constitution Prediction for on the other hand Spliced Proteins
55 Comparative Genomics equipment for the Prediction of Small RNA-Binding websites

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Extra info for Alternative pre-mRNA Splicing: Theory and Protocols

Sample text

It is likely that the speed of the polymerase activity is influenced by the chromatin structure, which indicates that the location of an exon in a certain chromatin region influences its regulation (Chapter 37 Kornblihtt) [33] An early event in transcription is formation of the 50 cap, which promotes spliceosome assembly [34]; similarly, the splicing of the last intron of a premRNA is coupled to the polyadenylation [34]. Finally, the splicing reaction deposits “exon junction complexes” just upstream of the newly formed exon–exon junction; these are necessary for an efficient RNA export, and they also enhance translation [35].

Currently, the most common genetic cause of death in children is spinal muscular atrophy (SMA). This deadly disease is caused by deletion of the SMN1 gene which, unfortunately, cannot be substituted by the almost identical SMN2 gene that is present in all patients. The difference between the two genes is a synonymous mutation that causes exon skipping in SMN2. Yet, the disease could be treated if the inclusion of this particular exon into the SMN2 mRNA could be promoted. Towards this aim, the regulation of the exon has been investigated in great detail (see Chapters 18 and 19 Singh), and this has led not only to the development of therapeutic approaches currently undergoing clinical trials [51], but also to new experimental approaches to alter splice site selection (see Chapter 45 Sch€ umperli).

Nature, 418, 222–228. R. (1982) Self-splicing RNA: autoexcision and autocyclization of the ribosomal RNA intervening sequence of Tetrahymena. Cell, 31, 147–157. , and Altman, S. (1983) The RNA moiety of ribonuclease P is the catalytic subunit of the enzyme. Cell, 35, 849–857. A. (2008) A structural 16 17 18 19 20 21 22 23 24 25 26 27 understanding of the dynamic ribosome machine. Nat. Rev. Mol. , 9, 242–253. L. (2007) Protein-free spliceosomal snRNAs catalyze a reaction that resembles the first step of splicing.

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Alternative pre-mRNA Splicing: Theory and Protocols by Stefan Stamm, Chris Smith, Reinhard Lührmann


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