Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

RNA immunoprecipitation (RIP)- qPCR (RIP-qPCR)

XW Xun Wu
YC Yi-Shing Lisa Cheng
MM Mathew Matthen
AY Angela Yoon
GS Gary K. Schwartz
SB Shashi Bala
AT Alison M. Taylor
FM Fatemeh Momen-Heravi
request Request a Protocol
ask Ask a question
Favorite

Cells were cross-linked and lysed in IP buffer supplemented with phosphatase/protease inhibitors and RNase inhibitor. Cell lysates were sonicated and 100 μg of total protein was incubated with Ago2 antibody or mouse IgG (non-specific control), the lysates were incubated overnight with either 10μg/ml of ChIP-grade anti-Ago2 or mouse IgG in 4 °C for 90 min, followed by addition of Protein A/G PLUS-Agarose beads and incubation for 80 min. After washing, the eluted RNA samples were further purified with TRI reagent and subjected to TaqMan MicroRNA Assay and qRT-PCR analysis to detect miR-34a-5p and MET, respectively. Data was normalized based on the total input.

Copyright and License information: The Author(s) ©2021
Open AccessThis article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. The images or other third party material in this article are included in the article's Creative Commons licence, unless indicated otherwise in a credit line to the material. If material is not included in the article's Creative Commons licence and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/. The Creative Commons Public Domain Dedication waiver (http://creativecommons.org/publicdomain/zero/1.0/) applies to the data made available in this article, unless otherwise stated in a credit line to the data.

Do you have any questions about this protocol?

Post your question to gather feedback from the community. We will also invite the authors of this article to respond.

0/150

tip Tips for asking effective questions

+ Description

Write a detailed description. Include all information that will help others answer your question including experimental processes, conditions, and relevant images.

post Post a Question
0 Q&A