Gold nanoparticles (gold NPs) modified with thiolated polyamide nucleic acids (PNA) carrying different charge moieties were prepared and characterized by UV-VIS spectroscopy, Photon Correlation Spectroscopy (PCS), and Transmission Electron Microscopy (TEM). The results demonstrated that the positively charged PNA modifier causes aggregation of the gold nanoparticle. By contrast, the gold nanoparticles modified with neutral and negatively charged PNAs are more stable towards aggregation. In all cases, the gold nanoparticles-bond PNA could not hybridize with their complementary DNAs. A new concept of DNA sequences determination based on gold nanoparticles aggregation induced by positively charged thiol-modified PNA was proposed. The PNA causes aggregation by neutralization of the negatively charge of the gold nanoparticles. If the PNA is first hybridized with DNA, the negatively-charge phosphate group of the DNA prevents PNA-induced aggregation in a sequence-specific manner. This concept has been validated and the technique has been used to determine single base mismatch in the DNA target of 9-13 base pair in length.