| Chapter title |
Metabolic Pulse-Chase RNA Labeling for pri-miRNA Processing Dynamics
|
|---|---|
| Chapter number | 3 |
| Book title |
miRNA Biogenesis
|
| Published in |
Methods in molecular biology, January 2018
|
| DOI | 10.1007/978-1-4939-8624-8_3 |
| Pubmed ID | |
| Book ISBNs |
978-1-4939-8623-1, 978-1-4939-8624-8
|
| Authors |
Annita Louloupi, Ulf Andersson Vang Ørom, Louloupi, Annita, Ørom, Ulf Andersson Vang |
| Abstract |
miRNA biogenesis is a multistep process starting with the cleavage of the primary miRNA transcript in the nucleus by the microprocessor complex. The pri-miRNA processing kinetics has a high impact on the final regulative role of the mature miRNAs on the expression of their target transcripts. Thus studying the in vivo kinetics of the miRNA biogenesis could give more insights into the contribution of each individual miRNA on regulation of gene expression. Here, we describe step by step a method to determine the processing kinetics of pri-miRNAs in vivo, using a pulse-chase approach that can be used in downstream applications such as qPCR or deep sequencing. We explain in detail the various aspects of this approach that can be applied to different mammalian cell types. The nature of this protocol allows the in vivo study of pri-miRNA processing kinetics in cells treated with different conditions, mutants, and/or cancer cell lines under physiological conditions. |
Mendeley readers
Geographical breakdown
| Country | Count | As % |
|---|---|---|
| Unknown | 7 | 100% |
Demographic breakdown
| Readers by professional status | Count | As % |
|---|---|---|
| Student > Ph. D. Student | 3 | 43% |
| Researcher | 1 | 14% |
| Professor > Associate Professor | 1 | 14% |
| Student > Master | 1 | 14% |
| Unknown | 1 | 14% |
| Readers by discipline | Count | As % |
|---|---|---|
| Biochemistry, Genetics and Molecular Biology | 3 | 43% |
| Agricultural and Biological Sciences | 1 | 14% |
| Medicine and Dentistry | 1 | 14% |
| Neuroscience | 1 | 14% |
| Unknown | 1 | 14% |