Analytical Sciences

Two-stage Nicking Enzyme Signal Amplification Combined with DNAzyme Amplification for the Detection of Bone Morphogenetic Protein 6 mRNA

Zhaohui HU, Yanhong ZHOU, Xiangtao XIE, Renjie JIANG, and Ningning LI

Department of Orthopaedics, Liuzhou People’s Hospital, 8 Wenchang Road, Liuzhou, 545006, Guangxi Province, P. R. China

Highly sensitive detection of bone morphogenetic protein 6 (BMP6) mRNA is essential to monitor bone regeneration in the regenerating defects. In this work, we proposed a quantitative approach based on two-stage nicking enzyme signal amplification (NESA) and DNAzyme amplification for highly sensitive detection of BMP6 mRNA. The two-stage NESA involves two templates and two-stage amplification reactions under isothermal conditions. The first template contains two repeat sequences that could hybridize to the target RNA, triggering an exponential amplification. The amplified product was a short single-stranded DNA with the same sequence as the target RNA. The single-stranded DNA can trigger another linear NESA and produces a large amount of horseradish peroxidase (HRP)-mimicking G-quadruplex DNAzyme. This proposed assay showed a quantitative analysis of BMP6 mRNA in a wide range from 1 fM to 100 nM with a detection limit of 0.01 fM.

J-STAGE: View this article in J-STAGE