| Chapter title |
Post-Transcriptional Gene Regulation
|
|---|---|
| Chapter number | 14 |
| Book title |
Post-Transcriptional Gene Regulation
|
| Published in |
Methods in molecular biology, January 2016
|
| DOI | 10.1007/978-1-4939-3067-8_14 |
| Pubmed ID | |
| Book ISBNs |
978-1-4939-3066-1, 978-1-4939-3067-8
|
| Authors |
Helwak, Aleksandra, Tollervey, David, Aleksandra Helwak, David Tollervey |
| Editors |
Erik Dassi |
| Abstract |
We present a detailed protocol for the experimental identification of miRNA-target RNA interaction sites using cross-linking, ligation, and sequencing of hybrids (CLASH). The basis of the technique is the purification of UV-stabilized Argonaute (AGO)-RNA complexes assembled in living cells, with subsequent ligation of AGO-associated RNA-RNA duplexes to form chimeric RNAs. Following cDNA synthesis, DNA library preparation and high-throughput sequencing, interacting RNA molecules are unambiguously identified as chimeric reads in bioinformatic analysis of sequencing data. CLASH potentially recovers any RNA duplex that is bound by RNA-binding protein, so modified approaches would be suitable for the identification of many other inter- and intramolecular RNA-RNA interactions. Since CLASH analysis is independent of bioinformatic predictions it allows the identification and analysis of RNA targeting rules in an unbiased way. |
X Demographics
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|---|---|---|
| Unknown | 1 | 100% |
Demographic breakdown
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|---|---|---|
| Members of the public | 1 | 100% |
Mendeley readers
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|---|---|---|
| United Kingdom | 1 | 3% |
| Unknown | 39 | 98% |
Demographic breakdown
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|---|---|---|
| Student > Ph. D. Student | 12 | 30% |
| Researcher | 7 | 18% |
| Student > Bachelor | 5 | 13% |
| Professor > Associate Professor | 4 | 10% |
| Student > Doctoral Student | 3 | 8% |
| Other | 7 | 18% |
| Unknown | 2 | 5% |
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|---|---|---|
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| Computer Science | 3 | 8% |
| Neuroscience | 1 | 3% |
| Engineering | 1 | 3% |
| Other | 0 | 0% |
| Unknown | 2 | 5% |