title: A novel magneto-DNA duplex probe for bacterial DNA detection based on exonuclease III-aided cycling amplification.
abstract: A novel magneto-DNA duplex probe for bacterial DNA detection based on exonuclease III (Exo-III) aided cycling amplification has been developed. This magneto-DNA duplex probe contains a partly hybrid fluorophore-modified capture probe and a fluorophore-modified signal probe with magnetic microparticle as carrier. In the presence of a perfectly matched target bacterial DNA, blunt 3'-terminus of the capture probe is formed, activating the Exo-III aided cycling amplification. Thus, Exo-III catalyzes the stepwise removal of mononucleotides from this terminus, releasing both fluorophore-modified signal probe, fluorescent dyes of the capture probe and target DNA. The released target DNA then starts a new cycle, while released fluorescent fragments are recovered with magnetic separation for fluorescence signal collection. This system exhibited sensitive detection of bacterial DNA, with a detection limit of 14 pM because of the unique cleavage function of Exo-III, high fluorescence intensity, and separating function of magneto-DNA duplex probes. Besides this sensitivity, this strategy exhibited excellent selectivity with mismatched bacterial DNA targets and other bacterial species targets and good applicability in real seawater samples, hence, this strategy could be potentially used for qualitative and quantitative analysis of bacteria.
authors.affiliation: Key Laboratory of Marine Environmental Corrosion and Bio-fouling, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China; University of the Chinese Academy of Science, 19 (Jia) Yuquan Road, Beijing 100039, China.
authors.forename: Yan
authors.initials: Y
authors.lastname: Zeng
authors.affiliation: Key Laboratory of Marine Environmental Corrosion and Bio-fouling, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China.
authors.forename: Yi
authors.initials: Y
authors.lastname: Wan
authors.affiliation: Key Laboratory of Marine Environmental Corrosion and Bio-fouling, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China. Electronic address: zhangdun@qdio.ac.cn.
authors.forename: Dun
authors.initials: D
authors.lastname: Zhang
authors.affiliation: Key Laboratory of Marine Environmental Corrosion and Bio-fouling, Institute of Oceanology, Chinese Academy of Sciences, 7 Nanhai Road, Qingdao 266071, China; University of the Chinese Academy of Science, 19 (Jia) Yuquan Road, Beijing 100039, China.
authors.forename: Peng
authors.initials: P
authors.lastname: Qi
mesh_terms: D015374:Biosensing Techniques; D015342:DNA Probes / Q000737:chemistry*; D004269:DNA, Bacterial / Q000737:chemistry / Q000302:isolation & purification*; D020564:Deltaproteobacteria / Q000737:chemistry*; D005090:Exodeoxyribonucleases / Q000737:chemistry*; D005456:Fluorescent Dyes; D057230:Limit of Detection; D059346:Magnets; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation*; D012623:Seawater / Q000382:microbiology; D013050:Spectrometry, Fluorescence
chemical_list: D015342:DNA Probes; D004269:DNA, Bacterial; D005456:Fluorescent Dyes; D005090:Exodeoxyribonucleases; C020133:exodeoxyribonuclease III
keywords: Bacterial DNA detection; Exonuclease III; Magnetic microparticle; Magneto-DNA duplex probe