A rapid room-temperature DNA amplification and detection strategy based on nicking endonuclease and catalyzed hairpin assembly

Abstract

A novel and rapid room-temperature DNA amplification strategy to simultaneously amplify and detect target HIV DNA was developed by coupling catalyzed hairpin assembly (CHA) with a “non-sequence dependent” nicking endonuclease.

Improving the Sensitivity of the miRNA Assay Coupled with the Mismatched Catalytic Hairpin Assembly Reaction by Optimization of Hairpin Annealing Conditions

“…The calibration curve obtained under the optimized conditions obeyed a double rectangular hyperbola equation with the formula ( R 2 0.9979), where a , b , c , and d were 2.03 × 10 6 , 0.12, 5.86 × 10 7 , and 365, respectively, and may be used to calculate the miRNA-141 concentration in the samples of interest. The curves with similar behaviors were previously reported for assays coupled with CHA/mCHA. ,, The analysis of the calibration curve showed that when the target concentration changed from 1 to 100 pM, the chemiluminescence intensity increased by 8 times, while in many of the miRNA assays reported previously, the signal value changed only by 3–4 times with changing the analyte concentration by 6–7 orders of magnitude. − …”

“…The curves with similar behaviors were previously reported for assays coupled with CHA/ mCHA. 28,32,33 The analysis of the calibration curve showed that when the target concentration changed from 1 to 100 pM, the chemiluminescence intensity increased by 8 times, while in many of the miRNA assays reported previously, the signal value changed only by 3−4 times with changing the analyte concentration by 6−7 orders of magnitude. 34−38 Study of Assay Specificity.…”

Recent advances of catalytic hairpin assembly and its application in bioimaging and biomedicine

“…In order to achieve genuine DNA self-replication, nicking endonuclease has been adopted in CHA-based strategies, to produce multiple copies of target DNA with the same sequence. Dong et al 47 coupled CHA with NtÁBsmAI to develop a rapid DNA amplification strategy for the HIV DNA target. The generated CHA product (H1-H2) would hybridize with another hairpin (H3) containing target DNA sequence, thus forming a Y shape DNA structure for the cleavage of NtÁBsmAI and producing massive replicas of target DNA.…”

Recent advances in cascade isothermal amplification techniques for ultra-sensitive nucleic acid detection

“…With the presence of the target, a pair of harpin probes formed a complex with a replica, which could be cyclically used as a new initiator of harpin probes. The self-replicating CHA has been successfully used to detect DNA and small molecules [ 157 , 158 ].…”