Rapid depletion of DIS3, EXOSC10 or XRN2 reveals the immediate impact of exoribonucleolysis on nuclear RNA metabolism and transcriptional control

Published: 4 February 2019| Version 6 | DOI: 10.17632/jyh2wdyb7z.6
Contributor:
Steven West

Description

This data represents uncropped western, co-immunoprecipitation and northern blots showing rapid depletion of EXOSC10 or DIS3 from human cells (westerns), effects of DIS3-AID tagging and EXOSC10 depletion of exosome integrity (Co-IP) and the effect of EXOSC10 loss on 5.8S rRNA processing (Northern).

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Steps to reproduce

All western blots were blocked for 1hr in 5% milk in PBS tween except for the AID blot which is blocked overnight in 10% milk in PBS tween. All primary antibodies were used at 1:1000 and secondary antibodies used at 1:2500. Antibody probing was for 1hr at room temperature in 2% milk in PBS tween. PBS tween was used to wash blots 3x in between primary and secondary antibodies and 3x before exposure. Blots were imaged using a biorad gel doc system. The Northern blot protocol is published in the accompanying manuscript.