title: Biosensing by Tandem Reactions of Structure Switching, Nucleolytic Digestion, and DNA Amplification of a DNA Assembly.
abstract: ϕ29 DNA polymerase (ϕ29DP) is able to carry out repetitive rounds of DNA synthesis using a circular DNA template by rolling circle amplification (RCA). It also has the ability to execute 3'-5' digestion of single-stranded but not double-stranded DNA. A biosensor engineering strategy is presented that takes advantage of these two properties of ϕ29DP coupled with structure-switching DNA aptamers. The design employs a DNA assembly made of a circular DNA template, a DNA aptamer, and a pre-primer. The DNA assembly is unable to undergo RCA in the absence of cognate target owing to the formation of duplex structures. The presence of the target, however, triggers a structure-switching event that causes nucleolytic conversion of the pre-primer by ϕ29DP into a mature primer to facilitate RCA. This method relays target detection by the aptamer to the production of massive DNA amplicons, giving rise to dramatically enhanced detection sensitivity.
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main Street West, Hamilton, ON, L8S 4K1 (Canada).
authors.forename: Meng
authors.initials: M
authors.lastname: Liu
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main Street West, Hamilton, ON, L8S 4K1 (Canada).
authors.forename: Wenqing
authors.initials: W
authors.lastname: Zhang
authors.affiliation: Department of Chemistry and Chemical Biology, McMaster University, 1280 Main Street West, Hamilton, ON, L8S 4K1 (Canada).
authors.forename: Qiang
authors.initials: Q
authors.lastname: Zhang
authors.affiliation: Department of Chemistry and Chemical Biology, McMaster University, 1280 Main Street West, Hamilton, ON, L8S 4K1 (Canada). brennanj@mcmaster.ca.
authors.forename: John D
authors.initials: JD
authors.lastname: Brennan
authors.affiliation: Department of Biochemistry and Biomedical Sciences, McMaster University, 1280 Main Street West, Hamilton, ON, L8S 4K1 (Canada). liying@mcmaster.ca.|Department of Chemistry and Chemical Biology, McMaster University, 1280 Main Street West, Hamilton, ON, L8S 4K1 (Canada). liying@mcmaster.ca.
authors.forename: Yingfu
authors.initials: Y
authors.lastname: Li
mesh_terms: D052157:Aptamers, Nucleotide / Q000737:chemistry* / Q000378:metabolism; D017103:Bacillus Phages / Q000201:enzymology; D015374:Biosensing Techniques / Q000379:methods*; D004247:DNA / Q000737:chemistry* / Q000378:metabolism; D004270:DNA, Circular / Q000737:chemistry / Q000378:metabolism; D004259:DNA-Directed DNA Polymerase / Q000378:metabolism; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*
chemical_list: D052157:Aptamers, Nucleotide; D004270:DNA, Circular; D004247:DNA; D004259:DNA-Directed DNA Polymerase
keywords: DNA amplification; aptamers; biosensors; rolling circle amplification; structure switching