Effects of Upconversion Nanoparticles on Polymerase Chain Reaction

Sang-Hyun Hwang,Su-Gyeong Im,Eun Jeong Lee,Yeon-Su Lee,Sang Soo Hah,Sang Jun Son,Geon Kook Lee,Do-Hoon Lee,Vu Thanh Cong,Richard C Willson

doi:10.1371/journal.pone.0073408

Sang-Hyun Hwang, Su-Gyeong Im + Show 8 more

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https://doi.org/10.1371/journal.pone.0073408

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Nanoparticles (NPs) are attractive materials owing to their physical and electrochemical properties, which make them extremely useful in diagnostic applications. Photon upconversion is the phenomenon where high-energy photons are emitted upon excitation of low-energy photons. Nucleic acids detection based on upconversion nanoparticles (UCNPs), which display a high signal-to-noise ratio and no photobleaching, has been widely applied. We evaluated whether UCNPs can improve polymerase chain reaction (PCR) specificity and affect PCR amplification. The effects of UCNPs with a diameter size of 40, 70, and 250 nm were evaluated using 3 PCR kits (AccuPower PCR PreMix, AmpliTaq Gold 360 Master Mix, and HotStarTaq Plus Master Mix) and 3 real-time PCR kits (AccuPower GreenStar qPCR PreMix, SYBR Green PCR Master Mix, and QuantiTect SYBR Green PCR Kit). Quantum dots were used for comparison with the UCNPs. In the presence of an appropriate concentration of UCNPs, PCR specificity was optimized. UCNPs of 40-nm size improved PCR specificity more effectively than did UCNPs sized 70 or 250 nm. As the size and concentrations of the UCNPs were increased, PCR amplification was more severely inhibited. At lower annealing temperatures (25°C–45°C), addition of the 40 nm UCNP (1 µg/µL) to the PCR reagent produced specific PCR products without nonspecific sequence amplification. Therefore, UCNPs of different sizes, with different DNA polymerases used in the commercial kits, showed different inhibitory effects on PCR amplification. These results demonstrate that optimization of UCNPs, added to reaction mixtures at appropriate concentrations, can improve PCR specificity. However, the mechanism underlining UCNPs effect on PCR remains unclear and will require further investigation.

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Polymerase chain reaction (PCR) techniques have been in widespread use in areas that require DNA/RNA detection, such as diagnosis and forensic analysis
The specificity of the PCR was improved at an appropriate concentration of upconversion nanoparticles (UCNPs) (Figure 2)
We demonstrated that the specificity of the PCR was effectively improved and no obvious effect on PCR amplification was seen at an appropriate concentration of the UCNPs of different sizes

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Polymerase chain reaction (PCR) techniques have been in widespread use in areas that require DNA/RNA detection, such as diagnosis and forensic analysis. Improving the PCR yield and specificity is important for developing more efficient molecular diagnostic tests. Some organic chemicals like formamide and amides, added to the reaction mixture, can enhance PCR amplification [1,2]. Nanoparticles (NPs) have become attractive materials owing to their physical and electrochemical properties, which make them extremely useful in the diagnostic applications for DNA [3], RNA [4], and protein [5]. Gold nanoparticles show tuned physical properties, including unique optical properties, robustness, and high surface areas, making them ideal candidates for developing biomarker platforms [7]

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