Open AccessTwo-stage Nicking Enzyme Signal Amplification Combined with DNAzyme Amplification for the Detection of Bone Morphogenetic Protein 6 mRNA
Author(s) -
Zhaohui Hu,
Yanhong Zhou,
Xing Xie,
Rong Jiang,
Ningning Li
Publication year - 2014
Publication title -
analytical sciences
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.392
H-Index - 73
eISSN - 1348-2246
pISSN - 0910-6340
DOI - 10.2116/analsci.30.1039
Subject(s) - deoxyribozyme , chemistry , microbiology and biotechnology , loop mediated isothermal amplification , horseradish peroxidase , dna , messenger rna , bone morphogenetic protein 6 , alkaline phosphatase , rna , bone morphogenetic protein , detection limit , rolling circle replication , biochemistry , enzyme , biology , bone morphogenetic protein 7 , polymerase , gene , chromatography
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.