| Chapter title |
Identification of mRNA-Interacting Factors by MS2-TRAP (MS2-Tagged RNA Affinity Purification)
|
|---|---|
| Chapter number | 2 |
| Book title |
RNA-Protein Complexes and Interactions
|
| Published in |
Methods in molecular biology, January 2016
|
| DOI | 10.1007/978-1-4939-3591-8_2 |
| Pubmed ID | |
| Book ISBNs |
978-1-4939-3589-5, 978-1-4939-3591-8
|
| Authors |
Je-Hyun Yoon, Myriam Gorospe, Yoon, Je-Hyun, Gorospe, Myriam |
| Abstract |
Posttranscriptional gene expression is governed by the interaction of mRNAs with vast families of RNA-binding proteins (RBPs) and noncoding (nc)RNAs. RBPs and ncRNAs jointly influence all aspects of posttranscriptional metabolism, including pre-mRNA splicing and maturation, mRNA transport, editing, stability, and translation. Given the impact of mRNA-interacting molecules on gene expression, there is great interest in identifying mRNA-binding factors comprehensively. Here, we provide a detailed protocol to tag mRNAs with MS2 hairpins and then affinity-purify trans-binding factors (RBPs, ncRNAs) associated with the MS2-tagged mRNA. This method, termed MS2-TRAP, permits the systematic characterization of ribonucleoprotein (RNP) complexes formed on a given mRNA of interest. We describe how to prepare the mRNA-MS2 expression vector, purify the MS2-tagged RNP complexes, and detect bound RNAs and RBPs, as well as variations of this methodology to address related questions of RNP biology. |
Mendeley readers
Geographical breakdown
| Country | Count | As % |
|---|---|---|
| Unknown | 79 | 100% |
Demographic breakdown
| Readers by professional status | Count | As % |
|---|---|---|
| Student > Ph. D. Student | 21 | 27% |
| Researcher | 11 | 14% |
| Student > Master | 7 | 9% |
| Student > Doctoral Student | 5 | 6% |
| Student > Bachelor | 5 | 6% |
| Other | 11 | 14% |
| Unknown | 19 | 24% |
| Readers by discipline | Count | As % |
|---|---|---|
| Biochemistry, Genetics and Molecular Biology | 31 | 39% |
| Agricultural and Biological Sciences | 15 | 19% |
| Neuroscience | 5 | 6% |
| Immunology and Microbiology | 5 | 6% |
| Chemistry | 2 | 3% |
| Other | 3 | 4% |
| Unknown | 18 | 23% |