title: In vitro selection of structure-switching, self-reporting aptamers.
abstract: We describe an innovative selection approach to generate self-reporting aptamers (SRAs) capable of converting target-binding events into fluorescence readout without requiring additional modification, optimization, or the use of DNA helper strands. These aptamers contain a DNAzyme moiety that is initially maintained in an inactive conformation. Upon binding to their target, the aptamers undergo a structural switch that activates the DNAzyme, such that the binding event can be reported through significantly enhanced fluorescence produced by a specific stacking interaction between the active-conformation DNAzyme and a small molecule dye, N-methylmesoporphyrin IX. We demonstrate a purely in vitro selection-based approach for obtaining SRAs that function in both buffer and complex mixtures such as blood serum; after 15 rounds of selection with a structured DNA library, we were able to isolate SRAs that possess low nanomolar affinity and strong specificity for thrombin. Given ongoing progress in the engineering and characterization of functional DNA/RNA molecules, strategies such as ours have the potential to enable rapid, efficient, and economical isolation of nucleic acid molecules with diverse functionalities.
authors.affiliation: Materials Department, University of California, Santa Barbara, CA 93106, USA.
authors.forename: Seung Soo
authors.initials: SS
authors.lastname: Oh
authors.forename: Kory
authors.initials: K
authors.lastname: Plakos
authors.forename: Xinhui
authors.initials: X
authors.lastname: Lou
authors.forename: Yi
authors.initials: Y
authors.lastname: Xiao
authors.forename: H Tom
authors.initials: HT
authors.lastname: Soh
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry* / Q000302:isolation & purification; D021881:DNA, Catalytic / Q000378:metabolism*; D005453:Fluorescence; D005456:Fluorescent Dyes; D015723:Gene Library; D006801:Humans; D008652:Mesoporphyrins; D009690:Nucleic Acid Conformation; D013917:Thrombin / Q000235:genetics
chemical_list: D052157:Aptamers, Nucleotide; D021881:DNA, Catalytic; D005456:Fluorescent Dyes; D008652:Mesoporphyrins; C065420:N-methylmesoporphyrin IX; D013917:Thrombin