Nicking enzyme amplification reaction

Nicking Enzyme Amplification Reaction (NEAR) is a method for in vitro DNA amplification like the polymerase chain reaction (PCR). NEAR is isothermal, replicating DNA at a constant temperature using a polymerase (and nicking enzyme) to exponentially amplify the DNA at a temperature range of 55 °C to 59 °C.

One disadvantage of PCR is that it consumes time uncoiling the double-stranded DNA with heat into single strands (a process called denaturation) . This leads to amplification times typically thirty minutes or more for significant production of amplified products.[1]

Potential advantages of NEAR over PCR are increased speed and lower energy requirements, characteristics that are shared with other isothermal amplification schemes.[2] A major disadvantage of NEAR relative to PCR is that production of nonspecific amplification products is a common issue with isothermal amplification reactions.[3]

The NEAR reaction uses naturally-occurring or engineered endonucleases that introduce a strand break on only one strand of a double-stranded DNA cleavage site. The ability of several of these enzymes to catalyze isothermal DNA amplification was disclosed but not claimed in the patents issued for the enzymes themselves.[4][5]

References

  1. ↑ Polymerase chain reaction
  2. ↑ Biosensors 2013, 3, 18-43; doi:10.3390/bios3010018
  3. ↑ Biochemistry. 2008 Sep 23;47(38):9987-99. doi: 10.1021/bi800746p.
  4. ↑ US Patent 6,191,267. February 20, 2001.
  5. ↑ US Patent 6,660,475. December 9, 2003


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