Impact of metal oxide nanoparticles on in vitro DNA amplification
Chun-Hui Gao,
Monika Mortimer,
Ming Zhang,
Patricia A. Holden,
Peng Cai,
Shan Wu,
Yuexing Xin,
Yichao Wu,
Qiaoyun Huang
Affiliations
Chun-Hui Gao
State Key Laboratory of Agricultural Microbiology, College of Resources and Environment, Huazhong Agricultural University, Wuhan, China
Monika Mortimer
Bren School of Environmental Science and Management, Earth Research Institute and University of California Center for the Environmental Implications of Nanotechnology (UC CEIN), University of California, Santa Barbara, Santa Barbara, CA, USA
Ming Zhang
State Key Laboratory of Agricultural Microbiology, College of Resources and Environment, Huazhong Agricultural University, Wuhan, China
Patricia A. Holden
Bren School of Environmental Science and Management, Earth Research Institute and University of California Center for the Environmental Implications of Nanotechnology (UC CEIN), University of California, Santa Barbara, Santa Barbara, CA, USA
Peng Cai
State Key Laboratory of Agricultural Microbiology, College of Resources and Environment, Huazhong Agricultural University, Wuhan, China
Shan Wu
State Key Laboratory of Agricultural Microbiology, College of Resources and Environment, Huazhong Agricultural University, Wuhan, China
Yuexing Xin
State Key Laboratory of Agricultural Microbiology, College of Resources and Environment, Huazhong Agricultural University, Wuhan, China
Yichao Wu
State Key Laboratory of Agricultural Microbiology, College of Resources and Environment, Huazhong Agricultural University, Wuhan, China
Qiaoyun Huang
State Key Laboratory of Agricultural Microbiology, College of Resources and Environment, Huazhong Agricultural University, Wuhan, China
Polymerase chain reaction (PCR) is used as an in vitro model system of DNA replication to assess the genotoxicity of nanoparticles (NPs). Prior results showed that several types of NPs inhibited PCR efficiency and increased amplicon error frequency. In this study, we examined the effects of various metal oxide NPs on inhibiting PCR, using high- vs. low-fidelity DNA polymerases; we also examined NP-induced DNA mutation bias at the single nucleotide level. The effects of seven major types of metal oxide NPs (Fe2O3, ZnO, CeO2, Fe3O4, Al2O3, CuO, and TiO2) on PCR replication via a low-fidelity DNA polymerase (Ex Taq) and a high-fidelity DNA polymerase (Phusion) were tested. The successfully amplified PCR products were subsequently sequenced using high-throughput amplicon sequencing. Using consistent proportions of NPs and DNA, we found that the effects of NPs on PCR yield differed depending on the DNA polymerase. Specifically, the efficiency of the high-fidelity DNA polymerase (Phusion) was significantly inhibited by NPs during PCR; such inhibition was not evident in reactions with Ex Taq. Amplicon sequencing showed that the overall error rate of NP-amended PCR was not significantly different from that of PCR without NPs (p > 0.05), and NPs did not introduce single nucleotide polymorphisms during PCR. Thus, overall, NPs inhibited PCR amplification in a DNA polymerase-specific manner, but mutations were not introduced in the process.
