Nicking endonuclease-mediated isothermal exponential amplification for double-stranded DNA detection

Abstract

Nucleic acid amplification and testing is routine demand in life science research and analytical chemistry. Polymerase chain reaction (PCR) is a widely employed DNA amplification technique using electrically powered thermal cycling equipment. The advent of isothermal amplification techniques circumvents the limitations of PCR for point-of-care testing (POCT). Here we have developed a novel method of exponential nicking endonuclease-mediated isothermal amplification for double-stranded DNA (dsDNA) detection. The new single-stranded DNA (ssDNA) target generation process occurs at a single temperature with next amplification reaction. The new ssDNA target copies are then exponentially amplified by alternating cycle strand displacement amplification (SDA). Therefore, it is a one-pot and isothermal strand displacement amplification method without the requirement of stepwise procedure, which greatly simplifies experimental procedure and decreases the probability of contamination of samples. With its advantages, it will be a useful tool to detect dsDNA in the field of POCT.