| Title: |
A universal method for the rapid isolation of all known classes of functional silencing small RNAs |
| Authors: |
Grentzinger, Thomas; Oberlin, Stefan; Schott, Gregory; Handler, Dominik; Svozil, Julia; Barragan-Borrero, Veronica; Humbert, Adeline; Duharcourt, Sandra; Brennecke, Julius; Voinnet, Olivier |
| Contributors: |
Institut Jacques Monod (IJM (UMR_7592)); Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité) |
| Source: |
ISSN: 0305-1048. |
| Publisher Information: |
CCSD; Oxford University Press |
| Publication Year: |
2020 |
| Subject Terms: |
[SDV]Life Sciences [q-bio] |
| Description: |
International audience ; Diverse classes of silencing small (s)RNAs operate via ARGONAUTE-family proteins within RNA-induced-silencing-complexes (RISCs). Here, we have streamlined various embodiments of a Q-sepharose-based RISC-purification method that relies on conserved biochemical properties of all ARGONAUTEs. We show, in multiple benchmarking assays, that the resulting 15-min benchtop extraction procedure allows simultaneous purification of all known classes of RISC-associated sRNAs without prior knowledge of the samples-intrinsic ARGONAUTE repertoires. Optimized under a user-friendly format, the method-coined 'TraPR' for Trans-kingdom, rapid, affordable Purification of RISCs-operates irrespectively of the organism, tissue, cell type or bio-fluid of interest , and scales to minute amounts of input material. The method is highly suited for direct profiling of silencing sRNAs, with TraPR-generated sequenc-ing libraries outperforming those obtained via gold-standard procedures that require immunoprecipita-tions and/or lengthy polyacrylamide gel-selection. TraPR considerably improves the quality and consistency of silencing sRNA sample preparation including from notoriously difficult-to-handle tissues/bio-fluids such as starchy storage roots or mammalian plasma, and regardless of RNA contaminants or RNA degradation status of samples. |
| Document Type: |
article in journal/newspaper |
| Language: |
English |
| DOI: |
10.1093/nar/gkaa472 |
| Availability: |
https://hal.science/hal-02990736; https://hal.science/hal-02990736v1/document; https://hal.science/hal-02990736v1/file/Grentzinger%20et%20al.%20-%202020%20-%20A%20universal%20method%20for%20the%20rapid%20isolation%20of%20all%20.pdf; https://doi.org/10.1093/nar/gkaa472 |
| Rights: |
info:eu-repo/semantics/OpenAccess |
| Accession Number: |
edsbas.FC67DDA1 |
| Database: |
BASE |