Library Science

A collection of readily reproducible techniques for the study of mRNA processing and metabolism. These cutting-edge techniques range from cotranscriptional processing events that occur while the mNA is engaged with elongating RNA polymerase II, to in vivo and in vitro splicing and its biochemical analysis, and alternative splicing. Additional methods cover mRNA export, the recovery and analysis of mRNP complexes, cytoplasmic translation, mRNA degradation in vivo and in vitro, and the controversial concept of nuclear translation. Each proven protocol is described in step-by-step detail and contains a background introduction outlining the principle behind the technique, lists of equipment and reagents, and tips on troubleshooting and avoiding know pitfalls.

Authors: Schoenberg, Daniel R.

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Table of Contents

1. Using Chromatin Immunoprecipitation to Map Cotranscriptional mRNA Processing in Saccharomyces cerevisiae (pp. 001-016)
   Keogh, Michael-Christopher; Buratowski, Stephen
2. Identifying PhosphoCTD-Associating Proteins (pp. 017-028)
   Phatnani, Hemali P.; Greenleaf, Arno L.
3. Imaging Alternative Splicing in Living Cells (pp. 029-046)
   Wagner, Eric J.; Baines, Andrea; Albrecht, Todd; Brazas, Robert M.; Garcia-Blanco, Mariano A.
4. Tobramycin Affinity Tag Purification of Spliceosomes (pp. 047-064)
   Hartmuth, Klaus; Vornlocher, Hans-Peter; Luhrmann, Reinhard
5. Using Single-Strand Conformational Polymorphism Gel Electrophoresis to Analyze Mutually Exclusive Alternative Splicing (pp. 065-074)
   Celotto, Alicia M.; Graveley, Brenton R.
6. Methods for the Analysis of Adenosine-to-Inosine Editing in RNA (pp. 075-084)
   Zhang, Zuo; Carmichael, Gordon G.
7. Assaying Nuclear Messenger RNA Export in Human Cells (pp. 085-092)
   Cullen, Bryan R.
8. RNA Unwinding Assay for DExD/H-Box RNA Helicases (pp. 093-102)
   Tseng-Rogenski, Stephanie S.; Chang, Tien-Hsien
9. Approaches for Monitoring Nuclear Translation (pp. 103-114)
   Iborra, Francisco J.; Jackson, Dean A.; Cook, Peter R.
10. Immunopurification and Analysis of Protein and RNA Components of mRNP in Mammalian Cells (pp. 115-124)
    Lejeune, Fabrice; Maquat, Lynne E.
11. Gene Expression Analysis of Messenger RNP Complexes (pp. 125-134)
    Penalva, Luiz O. F.; Tenenbaum, Scott A.; Keene, Jack D.
12. Using the (lambda)N Peptide to Tether Proteins to RNAs (pp. 135-154)
    Baron-Benhamou, Julie; Gehring, Niels H.; Kulozik, Andreas E.; Hentze, Matthias W.
13. An Efficient System for Cap- and Poly(A)-Dependent Translation In Vitro (pp. 155-170)
    Svitkin, Yuri V.; Sonenberg, Nahum
14. A Poly(A) Tail-Responsive In Vitro System for Cap- or IRES-Driven Translation From HeLa Cells (pp. 171-180)
    Thoma, Christian; Ostareck-Lederer, Antje; Hentze, Matthias W.
15. Assessing Messenger RNA Decapping in Cellular Extracts (pp. 181-192)
    Bergman, Naomi; Milone, Joseph; Bates, Elizabeth J.; Opyrchal, Mateusz; Bellofatto, Vivian; Wilusz, Jeffrey
16. Analysis of RNA Exonucleolytic Activities in Cellular Extracts (pp. 193-212)
    Mukherjee, Devi; Fritz, David T.; Kilpatrick, Walter J.; Gao, Min; Wilusz, Jeffrey
17. Application of Ligation-Mediated Reverse Transcription Polymerase Chain Reaction to the Identification of In Vivo Endonuclease-Generated Messenger RNA Decay Intermediates (pp. 213-222)
    Hanson, Mark N.; Schoenberg, Daniel R.
18. Biochemical Dissection of RNA Silencing in Plants (pp. 223-244)
    Tang, Guiliang; Zamore, Phillip D.
19. RNA Interference of mRNA Processing Factors in Drosophila S2 Cells (pp. 245-254)
    Celotto, Alicia M.; Graveley, Brenton R.
20. RNA Interference by Short Hairpin RNAs Expressed in Vertebrate Cells (pp. 255-266)
    Hannon, Gregory J.; Conklin, Douglas S.

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