Improve Research Reproducibility A Bio-protocol resource
Concise Method
tooltip

Quantitative Real-Time PCR

RL Robin A. Lu
AZ Amir A. Zeki
SR Sumati Ram-Mohan
NN Nhan Nguyen
YB Yan Bai
KC Kenneth Chmiel
SP Stevan Pecic
XA Xingbin Ai
RK Ramaswamy Krishnan
CG Chandra C. Ghosh
ask Ask a question
Favorite

Total RNA was extracted using the Quick-RNA Kit (Zymo Research, Irvine, CA). To transcribe the total RNA to cDNA, the qScript cDNA Synthesis Kit (Quantabio, Beverly, MA) was used. Next, we performed quantitative PCR (qPCR) reactions using the SYBR-green reaction mix (Bio-Rad, Hercules, CA) detected with the ABI 7500 Fast Real-Time PCR System (Thermo Fisher, Waltham, MA). Relative expression levels to ribosomal housekeeping controls (RPL-27) were determined using the comparative threshold method. Stable expression of RPL-27 was confirmed in all conditions. Total RNA for eotaxin-3 was extracted using Monarch Nucleic Acid Purification Kit (New England Biolabs, Ipswich, MA).

Copyright and License information:  ©2020 Lu, Zeki, Ram-Mohan, Nguyen, Bai, Chmiel, Pecic, Ai, Krishnan and Ghosh
This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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.

post Post a Question
0 Q&A