Selected article for: "HeLa cell and western blot"
Author: Nila Roy Choudhury; Gregory Heikel; Maryia Trubitsyna; Peter Kubik; Jakub Stanislaw Nowak; Shaun Webb; Sander Granneman; Christos Spanos; Juri Rappsilber; Alfredo Castello; Gracjan Michlewski
Title: RNA-binding activity of TRIM25 is mediated by its PRY/SPRY domain and is required for ubiquitination
  • Document date: 2017_10_9
    • ID: ifla4aix_100
    Snippet: Western blot analyses against T7 and DHX9 of pre-let-7a-1 pull downs with HeLa TRIM25 KO cell extracts overexpressing T7-TRIM25ΔPRY/SPRY or T7-TRIM25-PRY/SPR. Lanes 1 and 4 represent 4% (40 μ g) of the loading controls (input). Lanes 2 and 5 represent beads only controls. Lanes 3 and 6 show pre-let-7a-1 pull down. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. The copyright holder for this preprint (w.....
    Document: Western blot analyses against T7 and DHX9 of pre-let-7a-1 pull downs with HeLa TRIM25 KO cell extracts overexpressing T7-TRIM25ΔPRY/SPRY or T7-TRIM25-PRY/SPR. Lanes 1 and 4 represent 4% (40 μ g) of the loading controls (input). Lanes 2 and 5 represent beads only controls. Lanes 3 and 6 show pre-let-7a-1 pull down. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. The copyright holder for this preprint (which was not peer-reviewed) is the author/funder. show incubation of T7-TRIM25 with 6xHis-Ube2D3, UBE1 and ubiquitin for 5, 30 and 60 minutes, respectively. Lane 6 represents 60 minutes' reaction with T7-TRIM25, UBE1 and ubiquitin. Lanes 7, 8 and 9 show incubation of T7-TRIM25ΔRDB with 6xHis-Ube2D3, UBE1 and ubiquitin for 5, 30 and 60 minutes, respectively. Lane 10 represents 60 minutes' reaction with T7-TRIM25ΔRDB, UBE1 and ubiquitin. The proteins were analyzed by western blotting with anti-TRIM25 antibody. (c) Neither T7-TRIM25ΔRING nor T7-TRIM25K117R are efficiently ubiquitinated in vitro. Ubiquitination assays preformed on All rights reserved. No reuse allowed without permission.

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