title: A versatile biosensing system for DNA-related enzyme activity assay via the synthesis of silver nanoclusters using enzymatically-generated DNA as template.
abstract: In the present day, oligonucleotide-encapsulated silver clusters (DNA-AgNCs) have been widely applied into bio-analysis as a signal producer. Herein, we developed a novel method to synthesize DNA-AgNCs encapsulated by long-chain cytosine (C)-rich DNA. Such DNA was polymerized in a template-free way by terminal deoxynucleotidyl transferase (TdT). We demonstrated that TdT-polymerized long chain C-rich DNA can serve as an excellent template for AgNCs synthesis. Based on this novel synthesis strategy, we developed a label-free and turn-on fluorescence assay to detect TdT activity with ultralow limit of detection (LOD) of 0.0318 U and ultrahigh signal to background (S/B) of 46.7. Furthermore, our proposed method was extended to a versatile biosensing strategy for turn-on nucleases activity assay based on the enzyme-activated TdT polymerization. Two nucleases, EcoRI and ExoIII as model of endonuclease and exonuclease, respectively, have been detected with high selectivity and competitive low LOD of 0.0629 U and 0.00867 U, respectively. Our work demonstrates the feasibility of TdT polymerization-based DNA-AgNCs synthesis strategy as a versatile and potent biosensing platform to detect the activity of DNA-related enzymes.
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China.
authors.forename: Yijia
authors.initials: Y
authors.lastname: Yuan
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China.
authors.forename: Wenhua
authors.initials: W
authors.lastname: Li
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China.
authors.forename: Zhuoliang
authors.initials: Z
authors.lastname: Liu
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China. Electronic address: niezhou.hnu@gmail.com.
authors.forename: Zhou
authors.initials: Z
authors.lastname: Nie
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Huang
authors.affiliation: State Key Laboratory of Chemo/Biosensing and Chemometrics, College of Chemistry and Chemical Engineering, Hunan University, Changsha 410082, PR China.
authors.forename: Shouzhuo
authors.initials: S
authors.lastname: Yao
mesh_terms: D015374:Biosensing Techniques / Q000379:methods*; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D015246:Deoxyribonuclease EcoRI / Q000032:analysis* / Q000378:metabolism; D057075:Enzyme Assays / Q000379:methods*; D005090:Exodeoxyribonucleases / Q000032:analysis* / Q000378:metabolism; D057230:Limit of Detection; D049329:Nanostructures / Q000737:chemistry* / Q000648:ultrastructure; D058105:Polymerization; D012834:Silver / Q000737:chemistry*; D013050:Spectrometry, Fluorescence / Q000379:methods
chemical_list: D012834:Silver; D004247:DNA; D005090:Exodeoxyribonucleases; C020133:exodeoxyribonuclease III; D015246:Deoxyribonuclease EcoRI
keywords: DNA-templated synthesis; Label-free detection; Nuclease activity assay; Silver nanocluster; Terminal deoxynucleotidyl transferase