Related Experiment Video
Updated: Jul 20, 2026
High Sensitivity 5-hydroxymethylcytosine Detection in Balb/C Brain Tissue
Published on: February 1, 2011
Real-time MBDi-RPA using methyl-CpG binding protein 2: A real-time detection method for simple and rapid estimation
Mina Ishidoya1, Toshitsugu Fujita2, Sadatomo Tasaka1
1Department of Respiratory Medicine, Hirosaki University Graduate School of Medicine, 5 Zaifu-cho, Hirosaki, Aomori, 036-8562, Japan.
Background:
Analysis of CpG methylation is informative for cancer diagnosis. Previously, we developed a novel method to discriminate CpG methylation status in target DNA by blocking recombinase polymerase amplification (RPA), an isothermal DNA amplification technique, using methyl-CpG binding domain (MBD) protein 2 (MBD2). The method was named MBD protein interference-RPA (MBDi-RPA). In this study, MBDi-RPA was performed using methyl-CpG binding protein 2 (MeCP2), another MBD family protein, as the blocking agent.
Results:
MBDi-RPA using MeCP2 detected low levels of CpG methylation, showing that it had higher sensitivity than MBDi-RPA using MBD2. We also developed real-time RPA, which enabled rapid analysis of DNA amplification without the need for laborious agarose gel electrophoresis and used it in combination with MBDi-RPA. We termed this method real-time MBDi-RPA. The method using MeCP2 could determine the abundance ratio of CpG-methylated target DNA simply and rapidly, although highly sensitive detection was challenging.
Significance And Novelty:
Real-time MBDi-RPA using MeCP2 could be potentially useful for estimating CpG methylation status in target DNA prior to more detailed analyses.
