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Issue 6, 2018
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Technical aspects of nicking enzyme assisted amplification

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Abstract

Nicking enzyme assisted amplification (NEAA) is an extremely rapid method for molecular diagnosis. However, this technology is not widely applied for real sample analysis because the overproduced non-specific products limit its sensitivity and raise the threshold of detection methods. Here, we have found that the non-specific amplification is mainly caused by the coexistence of Bst polymerase, nicking primers and dNTP. The highly active nicking enzyme directs and accelerates the non-specific amplification in a way which favors nicking. To suppress the non-specific amplification, the nicking enzyme concentration, reaction temperature, and magnesium ion concentration are optimized. The compatibility of Bst polymerase with the concentration of the monovalent cation is also crucial. Besides, the sensitivity could be enhanced by shortening the target sequences and priming the 3′ end of the target.

Graphical abstract: Technical aspects of nicking enzyme assisted amplification

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Publication details

The article was received on 17 Dec 2017, accepted on 28 Jan 2018 and first published on 29 Jan 2018


Article type: Paper
DOI: 10.1039/C7AN02037F
Citation: Analyst, 2018,143, 1444-1453
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    Technical aspects of nicking enzyme assisted amplification

    L. Wang, C. Qian, H. Wu, W. Qian, R. Wang and J. Wu, Analyst, 2018, 143, 1444
    DOI: 10.1039/C7AN02037F

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