Effects of Single-Walled Carbon Nanotube on Polymerase Chain Reaction
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Effects of Single-Walled Carbon Nanotube on Polymerase Chain Reaction Daxiang Cui, Furong Tian,Yong Kong, Cengiz S. Ozkan1, Igor Titushikin and Huajian Gao Max Planck Institute for Metals Research, Heisenbergstrasse 3, 70569 Stuttgart, Germany 1 Department of Mechanical Engineering, University of California, Riverside, CA 92521-0425, USA ABSTRACT Polymerase Chain Reaction (PCR) is a complicated gene amplification process using Mg2+ as an assistant factor of Taq enzyme. Influence of single-walled carbon nanotube on that reaction was investigated with scanning electron microscope (SEM), transmission electron microscope (TEM) and quantitative PCR product analysis. Results showed that adding single-walled carbon nanotubes in the reaction liquid with a concentration of less than 3 µg/µl increase the amounts of PCR products. But more carbon nanotubes cause a great reduction of the PCR products. Similar effects were observed in the reaction without Mg2+. Possible reason could be the aggregation of reaction components caused by small amount of additive carbon nanotubes, which increases reaction probability. Our observations suggest that small amount of carbon nanotube could be used as an assistant factor to improve the PCR reaction.
INTRODUCTION Polymerase Chain Reaction (PCR) was invented by Kary Banks Mullis in 1985, also called gene amplification technology. This technique can increase the amounts of target gene fragments by a magnification of several millions [1]. The reaction consists of a three-step recycling course: denaturation, anneal and extension. Its reaction components include 10×PCR buffer, dNTPs mixture, Mg2+, DNA template and Taq enzyme. In the reaction course, Mg2+ is a very important component to improve the activity of Taq enzyme. With less or without Mg2+ the reaction efficiency is very low. Single-walled carbon nanotubes (SWCNTs) possess excellent physical and chemical properties, and can interact with nucleic acids and protein [2-3] showing very good bio-compatibility. In our present study, effects of SWCNT on PCR reaction with or without Mg2+ ion was investigated, with the aim of exploring the potential interaction mechanism between carbon nanotube and target gene fragments as well as Taq enzyme.
EXPERIMENT DETAILS SWCNTs with diameter of 2nm (from Carbon Nanotechnologies Inc.) were dissolved in the ion-free water, with final concentration of 10mg/ml. Gene vector with brcaa1 (breast cancer associated antigen 1gene, AF208045) was dissolved in the ion-free water, with final concentration of 0.1µg/µl. PCR primers were diluted into 10pmol/ µl. The upstream primer sequence is 5’-CGC TTA ATT AAA CAT ATG ACC AGA GTG AAA GAT GCT CAG-3’ and the downstream primer sequence 5’- TTA GTT AGT TAC CGG ATC CCT TAA CTC CAT TTG TAA ACT TTG G –3’. Target gene fragment is 410bp in length.
Downloaded from https://www.cambridge.org/core. Access paid by the UCSB Libraries, on 01 Apr 2018 at 12:47:37, subject to the Cambridge Core terms of use, available at https://www.cambridge.org/core/terms. https://doi.org/10.1
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