title: Thioaromatic DNA monolayers for target-amplification-free electrochemical sensing of environmental pathogenic bacteria.
abstract: Genosensing technology has mostly based on mixed self-assembled monolayers (SAMs) of thiol-modified oligonucleotides and alkanethiols on gold surfaces. However, the typical backfilling approach, which incorporates the alkanethiol in a second step, gives rise to a heterogeneous distribution of oligonucleotide probes on the surface, negatively affecting to both hybridization efficiency and surface stability. Despite aromatic thiols present a remarkably different behavior from alkanethiols, with higher rigidity and stronger intermolecular interactions, they have been scarcely explored for the fabrication of DNA sensing platforms. We have investigated different approaches involving SAMs of aromatic thiols, namely p-mercaptobenzoic acid (p-MBA) and p-aminothiophenol (p-ATP), to yield DNA sensing layers for sequence-specific detection of target oligonucleotides. The studied monolayers were evaluated by DNA surface coverage and further information was obtained by determining their functionality in a sandwich hybridization assay with enzymatic amplification of the electrochemical read-out. The insertion of thiol-oligonucleotides into p-ATP monolayers previously oxidized, and the covalent binding of amino-oligonucleotides to pure p-MBA monolayers give rise to increased storage stability and better analytical performance. The quantification of RNA from Legionella pneumophila cellular lysates was successfully performed, illustrating the usefulness of these sensing architectures for detecting pathogenic bacteria.
authors.affiliation: Departamento de Química Física y Analítica, Universidad de Oviedo, 33006 Oviedo, Spain.
authors.forename: Rebeca
authors.initials: R
authors.lastname: Miranda-Castro
authors.affiliation: Departamento de Química Física y Analítica, Universidad de Oviedo, 33006 Oviedo, Spain.
authors.forename: Raquel
authors.initials: R
authors.lastname: Sánchez-Salcedo
authors.affiliation: Departamento de Química Física y Analítica, Universidad de Oviedo, 33006 Oviedo, Spain.
authors.forename: Beatriz
authors.initials: B
authors.lastname: Suárez-Álvarez
authors.affiliation: Departamento de Química Física y Analítica, Universidad de Oviedo, 33006 Oviedo, Spain.
authors.forename: Noemí
authors.initials: N
authors.lastname: de-Los-Santos-Álvarez
authors.affiliation: Departamento de Química Física y Analítica, Universidad de Oviedo, 33006 Oviedo, Spain.
authors.forename: Arturo J
authors.initials: AJ
authors.lastname: Miranda-Ordieres
authors.affiliation: Departamento de Química Física y Analítica, Universidad de Oviedo, 33006 Oviedo, Spain. Electronic address: mjlc@uniovi.es.
authors.forename: María
authors.initials: M
authors.lastname: Jesús Lobo-Castañón
mesh_terms: D000814:Aniline Compounds / Q000737:chemistry*; D001565:Benzoates / Q000737:chemistry*; D015374:Biosensing Techniques / Q000379:methods*; D055664:Electrochemical Techniques / Q000379:methods; D006046:Gold / Q000737:chemistry; D006801:Humans; D061925:Immobilized Nucleic Acids / Q000737:chemistry*; D016952:Legionella pneumophila / Q000302:isolation & purification*; D007877:Legionnaires' Disease / Q000382:microbiology*; D057230:Limit of Detection; D009693:Nucleic Acid Hybridization / Q000379:methods; D012329:RNA, Bacterial / Q000032:analysis*; D012335:RNA, Ribosomal / Q000032:analysis; D013438:Sulfhydryl Compounds / Q000737:chemistry*; D013499:Surface Properties; D014871:Water Microbiology
chemical_list: D000814:Aniline Compounds; D001565:Benzoates; D061925:Immobilized Nucleic Acids; D012329:RNA, Bacterial; D012335:RNA, Ribosomal; D013438:Sulfhydryl Compounds; C013594:4-mercaptobenzoate; D006046:Gold; C064316:4-aminothiophenol
keywords: Backfilling method; Bacterial RNA detection; DNA sensing surfaces; Inserting method; Legionella pneumophila; Thioaromatic self-assembled monolayers