科研之友
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摘要
Polymerase chain reaction (PCR)-enhancing effect of gold nanoparticles (GNPs) has been reported earlier. Two opinions exist; one suggests that the enhancement arises due to an enrichment of heat transfer by high-conductivity, high-heat capacity GNPs while the other indicates that the PCR-enhancing effect of GNPs is mediated by surface interactions of the GNPs rather than by heat-transfer enhancement. In this context the present study aimed at deciphering the role of GNP in regulating the yield of PCR. GNPs similar to 11 nm in size were synthesized and characterized by transmission electron microscopy, atomic force microscopy, dynamic light scattering and UV-visible spectroscopy. An increase in the yield of PCR product was observed initially with the addition of increasing concentrations of GNPs, reaching a maximum value and declining thereafter. To monitor the influence of GNPs on PCR, GNPs interacting with different reacting components of PCR were monitored by UV-visible spectroscopy, fluorescence quenching, circular dichroism spectroscopy and AFM. UV-visible spectroscopy and circular dichroism studies showed that GNPs had a greater affinity for Taq DNA polymerase compared to the primer or DNA template. Fluorescence quenching studies showed notable quenching of the spectra of Taq DNA polymerase in the presence of GNPs, indicating a direct interaction of the GNPs with the Taq chromophores. Taq-conjugated gold nanoparticles were visualized by AFM. A CD melting study showed that the inversion temperature of Taq polymerase increased from 73 degrees C to 81 degrees C in presence of GNP. The enhancement of PCR yield by GNPs could be attributed to the greater affinity and thermodynamic stability of GNPs for Taq DNA polymerase compared to the primer or DNA template. These findings shed light toward the trafficking power of nanoparticles in various enzymatic reactions to regulate the yield of the product.
- 出版日期2013
