title: KF polymerase-based fluorescence aptasensor for the label-free adenosine detection.
abstract: We have developed a simple, inexpensive, and label-free method for the selective detection of adenosine. Klenow fragment polymerase (KF polymerase) is a commonly-used 5' to 3' DNA polymerase, it also has 3' to 5' exonuclease activity that can digest single-stranded DNA. An adenosine binding DNA aptamer was employed, the aptamer was split into two pieces of single-stranded DNA (aptamer-A1 + aptamer-A2). Without the addition of adenosine, aptamer-A1 and aptamer-A2 existed as single-stranded DNA which could be efficiently degraded by the exonuclease activity of KF polymerase. Much reduced background fluorescence was obtained when SYBR Green dye was added. However, in the presence of adenosine, aptamer-A1 and aptamer-A2 bound to adenosine, and hybridization of the complementary sequences resulted in the formation of a duplex DNA structure, which could initiate DNA polymerization. The addition of SYBR Green dye resulted in a very high fluorescence enhancement, which could be used for the quantification of adenosine.
authors.affiliation: State Key Laboratory of Electroanalytical Chemistry, Changchun Institute of Applied Chemistry, Chinese Academy of Sciences, Changchun, Jilin 130022, PR China.
authors.forename: Dongli
authors.initials: D
authors.lastname: Liao
authors.forename: Huping
authors.initials: H
authors.lastname: Jiao
authors.forename: Bin
authors.initials: B
authors.lastname: Wang
authors.forename: Quan
authors.initials: Q
authors.lastname: Lin
authors.forename: Cong
authors.initials: C
authors.lastname: Yu
mesh_terms: D000241:Adenosine / Q000032:analysis* / Q000737:chemistry; D052157:Aptamers, Nucleotide / Q000737:chemistry; D052160:Benzothiazoles; D015374:Biosensing Techniques / Q000379:methods*; D004256:DNA Polymerase I / Q000737:chemistry*; D004277:DNA, Single-Stranded / Q000737:chemistry*; D003959:Diamines; D004591:Electrophoresis, Polyacrylamide Gel / Q000379:methods; D009693:Nucleic Acid Hybridization / Q000235:genetics; D009930:Organic Chemicals; D011804:Quinolines; D013194:Staining and Labeling
chemical_list: D052157:Aptamers, Nucleotide; D052160:Benzothiazoles; D004277:DNA, Single-Stranded; D003959:Diamines; D009930:Organic Chemicals; D011804:Quinolines; C098022:SYBR Green I; D004256:DNA Polymerase I; D000241:Adenosine