Nickase-dependent isothermal DNA amplification

We developed a nicking endonuclease dependent DNA amplification (NDA), using Nt.BstNBI to catalyze single-stranded nick on double-stranded DNA, and Bst DNA polymerase to make extension while sealing the nick and displacing the downstream strand. The displaced single-stranded DNA thereby serves as template for primers hybridization and extension, resulting in exponential synthesis of target DNA under isothermal condition. Over 105 folds target DNA amplification can be achieved in 30 minutes, generating DNA product suitable for both diagnosis and DNA cloning. This NDA strategy does not require thermal cycling or prerequisite nucleotides modification, making it suitable for application in the field and at the point-of-care.