Abstract
Gold nanoparticles (AuNPs) have been reported to induce faster heat transfer of polymerase chain reaction (PCR) solution to enhance PCR efficiency. AuNPs can also increase the specificity of PCR by functioning analogously to single-stranded DNA binding protein (SSB). However, the simple structure of AuNPs makes it difficult to determine how AuNPs affect PCR efficiency and specificity. This study aimed to elucidate the effect of AuNPs on PCR efficiency by altering the denaturation times and annealing temperatures. In addition, comparative study of the effect of AuNPs on PCR inhibition caused by two competitive primers allowed investigation of the mechanisms of AuNP-enhanced PCR accuracy. Finally, heat-treated salmon sperm DNA was used to evaluate whether AuNPs could eliminate the inhibitory PCR effect caused by DNA impurities. This study demonstrated that enhanced thermal conductivity by AuNPs was the main mechanism for increased PCR efficiency and specificity. AuNPs promoted efficient double-stranded DNA template unwinding and dissociation between mismatched primers, DNA fragments and template, and enhanced PCR efficiency and specificity simultaneously. Thus, this significant finding suggests the future use of AuNP-assisted PCR in different fields, especially in rapid clinical diagnosis and screening by PCR.
Highlights
Gold nanoparticles (AuNPs) have been reported to induce faster heat transfer of polymerase chain reaction (PCR) solution to enhance PCR efficiency
Inc. (China), 2×PCR Master Mix was purchased from Tiangen (China), salmon sperm DNA was purchased from Sigma (USA), and the primers were synthesized by Sangon Biological Engineering Technology and Services (China)
At lower concentrations, or high concentrations of the initial DNA template (2.5 and 25 ng/μL), the effect of AuNPs was not observed. These results indicate that AuNPs could significantly enhance PCR efficiency only at appropriate DNA template concentrations
Summary
Gold nanoparticles (AuNPs) have been reported to induce faster heat transfer of polymerase chain reaction (PCR) solution to enhance PCR efficiency. AuNPs can increase the specificity of PCR by functioning analogously to single-stranded DNA binding protein (SSB). AuNPs promoted efficient double-stranded DNA template unwinding and dissociation between mismatched primers, DNA fragments and template, and enhanced PCR efficiency and specificity simultaneously. This significant finding suggests the future use of AuNP-assisted PCR in different fields, especially in rapid clinical diagnosis and screening by PCR. By increasing the annealing temperature to reduce the mispairings between primers and DNA templates, the amplification specificity of PCR can be improved. The specific binding of AuNPs to ssDNA template reduces mismatch between primers and DNA template, thereby increasing the specificity of PCR amplification. The use of two different mechanisms to explain similar biological effects requires further study
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