The isothermal amplification detection of double-stranded DNA based on a double-stranded fluorescence probe

Biosensors & Bioelectronics
Chao ShiCuiping Ma

Abstract

Here we have developed a novel method of isothermal amplification detection of double-stranded DNA (dsDNA) based on double-stranded fluorescence probe (ds-probe). Target dsDNA repeatedly generated single-stranded DNA (ssDNA) with polymerase and nicking enzyme. The ds-probe as a primer hybridized with ssDNA and extended to its 5'-end. The displaced ssDNA served as a new detection target to initiate above-described reaction. Meanwhile, the extended ds-probe could dynamically dissociate from ssDNA and self-hybridize, converting into a turn-back structure to initiate another amplification reaction. In particular, the ds-probe played a key role in the entire experimental process, which not only was as a primer but also produced the fluorescent signal by an extension and displacement reaction. Our method could detect the pBluescript II KS(+) plasmid with a detection limit of 2.3 amol, and it was also verified to exhibit a high specificity, even one-base mismatch. Overall, it was a true isothermal dsDNA detection strategy with a strongly anti-jamming capacity and one-pot, only requiring one ds-probe, which greatly reduced the cost and the probability of contamination. With its advantages, the approach of dsDNA detection will offer a p...Continue Reading

References

Apr 11, 1992·Nucleic Acids Research·G T WalkerD P Malinowski
Apr 17, 2004·Nucleic Acids Research·Jinping ChengQingge Li
Aug 23, 2005·Clinica Chimica Acta; International Journal of Clinical Chemistry·Salvatore A E MarrasFred Russell Kramer
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Jan 22, 2011·Bioanalysis·Joonyul Kim, Christopher J Easley
Mar 10, 2011·Lab on a Chip·Peter J Asiello, Antje J Baeumner

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Citations

Jun 3, 2017·European Journal of Clinical Microbiology & Infectious Diseases : Official Publication of the European Society of Clinical Microbiology·P MaffertH Abaibou

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