Intermolecular and Intramolecular Priming Co-directed Synergistic Multi-strand Displacement Amplification Empowers Ultrasensitive Determination of microRNAs

Abstract

The construction of isothermal nucleic acid amplification systems with extremely high signal amplification abilities is of great importance for biomarker detection and disease diagnosis. Herein, for the first time, we proposed an intermolecular and intramolecular priming co-directed synergistic multi-strand displacement amplification (SM-SDA) system for microRNA (miRNA) detection. Strategically, the SM-SDA system is made of a single multi-functionalized hairpin probe (MF-HP) that is engineered with a long stem and endowed with target complementation, configuration transformation, enzyme recognition, and signal reporting abilities. The presence of a specific target, the model of miRNA-21, to react with MF-HP would cause an intermolecular priming-directed strand replication in parallel with an intramolecular priming-directed strand replication. The codirected priming pattern results in the occurrence of SM-SDA containing a target-induced cyclical strand displacement amplification (T-CSDA), a target analogue primer (TAP)-induced CSDA (TAP-CSDA), and a nicked trigger primer (NTP)-induced CSDA (NTP-CSDA). The resulting multiple circuits with a synergetic signal amplification capacity remarkably enhance the target miRNA response, which has not only improved the sensitivity for qualitative and quantitative detection of miRNA but also realized the analysis of target miRNA from real clinical samples. As a proof-of-concept study, this simple but attractive isothermal signal amplification system holds a great potential for molecular diagnosis of diseases and will stimulate interest, new ideas, and discoveries in this fascinating field.

A DNA polymerase-powered self-propelled DNA walking strategy for one-step, amplified and dual-signal electrochemical target detection

“…Alternatively, some DNA-based isothermal amplication strategies, for example, rolling circle amplication (RCA), loopmediated isothermal amplication (LAMP), helicase dependent amplication (HDA), recombinase polymerase amplication (RPA), and strand displacement amplication (SDA), are popular for sensitivity improvement. [17][18][19][20] However, these strategies are usually based on the combined use of several nucleases, increasing the assay cost and the possibility of non-specic amplication. Also, these signal amplication means are more suitable for nucleic acid analysis by optical methods.…”

An allosteric palindromic hairpin probe based dual-mode interactive strand displacement amplification enables robust miRNA biosensing

“…MicroRNAs (miRNAs) are a class of endogenous small noncoding RNAs that regulate gene expression 1 The dysregulated expression of specific miRNAs, especially those circulating miRNAs expressed in serum, plasma, tears, and other body fluids, has been associated with various human diseases, including cancer, highlighting their potential as valuable biomarkers for cancer diagnosis and therapy. 2,3 For instance, miRNA-155 exhibits high expression in blood in breast cancer. 4 Considering that early diagnosis always represents the most promising strategies in combating cancer, it is crucial to develop new methods towards miRNA detection.…”