title: Label-free monitoring of DNA polymerase activity based on a thrombin-binding aptamer G-quadruplex.
abstract: We have developed a label-free assay for the detection of DNA polymerase activity based on a thrombin-binding aptamer (TBA) G-quadruplex. In the presence of DNA polymerase, the 3'-OH termini of the hairpin substrate are immediately elongated to replace the TBA, which can be recognized quickly by the ThT dye and results in an increase of fluorescence. This method is highly sensitive with a detection limit of 0.1 U/mL. It is simple and cost-effective without any requirement of labeling with a fluorophore-quencher pair. Furthermore, the proposed method can also be applied to analyze the inhibition of DNA polymerase, which clearly indicates that the proposed method can be applied for screening of potential DNA polymerase inhibitors.
authors: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha 410013, China.
authors: Jing
authors: J
authors: Wang
authors: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha 410013, China.
authors: Haisheng
authors: H
authors: Liu
authors: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha 410013, China. Electronic address: macb2012@csu.edu.cn.
authors: Changbei
authors: C
authors: Ma
authors: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha 410013, China.
authors: Jun
authors: J
authors: Wang
authors: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha 410013, China.
authors: Linxiu
authors: L
authors: Zhong
authors: State Key Laboratory of Medical Genetics, School of Life Sciences, Central South University, Changsha 410013, China.
authors: Kefeng
authors: K
authors: Wu
mesh_terms: D052157:Aptamers, Nucleotide / Q000378:metabolism*; D001483:Base Sequence; D052160:Benzothiazoles; D004256:DNA Polymerase I / Q000378:metabolism; D015342:DNA Probes / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism*; D005453:Fluorescence; D054856:G-Quadruplexes; D013194:Staining and Labeling; D013844:Thiazoles / Q000378:metabolism; D013997:Time Factors
chemical_list: D052157:Aptamers, Nucleotide; D052160:Benzothiazoles; D015342:DNA Probes; D013844:Thiazoles; C081676:thrombin aptamer; C009462:thioflavin T; D004256:DNA Polymerase I; D004259:DNA-Directed DNA Polymerase
keywords: DNA polymerase; Label-free; Thioflavin T; Thrombin-binding aptamer G-quadruplex