Nicking and Extension Amplification Reaction for the Exponential Amplification of Nucleic Acids

A method of amplifying a target nucleotide sequence, the method comprising:

• (a) combining a target nucleic acid comprising the target nucleotide sequence with an amplification reagent mixture comprising:
• i. a polymerase,
• ii. one or more nicking enzymes,
• iii. a first oligonucleotide comprising a 5′ portion that comprises a nicking site that is non-complementary to the target nucleotide sequence and a 3′ portion that hybridizes to the target nucleotide sequence, and
• iv. a second oligonucleotide comprising a 5′ portion that comprises a nicking site that is non-complementary to the target nucleotide sequence and a 3′ portion that hybridizes to the target nucleotide sequence, and
• (b) subjecting the reaction mixture formed by the step of combining to essentially isothermal conditions to amplify the target nucleotide sequence, and
• (c) detecting the amplified target nucleotide sequence.

A system configured to amplify a target nucleotide sequence under essentially isothermal conditions and detect the amplified target nucleotide sequence, the system comprising an amplification reagent mixture comprising:

• i. a polymerase,
• ii. one or more nicking enzymes,
• iii. a first oligonucleotide comprising a 5′ portion that comprises a nicking site that is non-complementary to the target nucleotide sequence and a 3′ portion that hybridizes to the target nucleotide sequence, and
• iv. a second oligonucleotide comprising a 5′ portion that comprises a nicking site that is non-complementary to the target nucleotide sequence and a 3′ portion that hybridizes to the target nucleotide sequence.