Author, as appears in the article.: AL-Madhagi, Sallam; O'Sullivan, Ciara K.; Prodromidis, Mamas I.; Katakis, Ioanis;
Department: Enginyeria Química
URV's Author/s: Katakis, Ioanis / O'SULLIVAN, CIARA KATHLEEN
Keywords: Recombinase polymerase amplification Modified primers Karlodinium-veneficum Karlodinium armiger sequence in seawater Isothermal amplification Genomic dna Functionalized gold nanoparticles Electrochemical detection Dna detection Biosensors
Abstract: A reagent-less DNA sensor has been developed exploiting a combination of gold nanoparticles, modified primers, and isothermal amplification. It is applied to the determination ofKarlodinium armiger, a toxic microalgae, as a model analyte to demonstrate this generic platform. Colloidal gold nanoparticles with an average diameter of 14 +/- 0.87 nm were modified with a mixed self-assembled monolayer of thiolated 33-mer DNA probes and (6-mercaptohexyl) ferrocene. Modified primers, exploiting a C3 spacer between the primer-binding site and an engineered single-stranded tail, were used in an isothermal recombinase polymerase amplification reaction to produce an amplicon by two single-stranded tails. These tails were designed to be complementary to a gold electrode tethered capture oligo probe, and an oligo probe immobilized on the gold nanoparticles, respectively. The time required for hybridization of the target tailed DNA with the surface immobilized probe and reporter probe immobilized on AuNPs was optimized and reduced to 10 min, in both cases. Amplification time was further optimized to be 40 min to ensure the maximum signal. Under optimal conditions, the limit of detection was found to be 1.6 fM of target dsDNA. Finally, the developed biosensor was successfully applied to the detection of genomic DNA extracted from a seawater sample that had been spiked with K. armiger cells. The demonstrated generic electrochemical genosensor can be exploited for the detection of any DNA sequence and ongoing work is moving towards an integrated system for use at the point-of-need.
Thematic Areas: Zootecnia / recursos pesqueiros Química Materiais Interdisciplinar Geociências Engenharias iv Engenharias ii Ciências biológicas i Ciências ambientais Chemistry, analytical Biotecnología Biodiversidade Astronomia / física Analytical chemistry
licence for use: https://creativecommons.org/licenses/by/3.0/es/
Author's mail: ioanis.katakis@urv.cat
Author identifier: 0000-0003-4259-7098
Record's date: 2023-02-19
Papper version: info:eu-repo/semantics/acceptedVersion
Link to the original source: https://link.springer.com/article/10.1007/s00604-021-04771-8
Licence document URL: http://repositori.urv.cat/ca/proteccio-de-dades/
Papper original source: Microchimica Acta. 188 (4):
APA: AL-Madhagi, Sallam; O'Sullivan, Ciara K.; Prodromidis, Mamas I.; Katakis, Ioanis; (2021). Combination of ferrocene decorated gold nanoparticles and engineered primers for the direct reagentless determination of isothermally amplified DNA. Microchimica Acta, 188(4), -. DOI: 10.1007/s00604-021-04771-8
Article's DOI: 10.1007/s00604-021-04771-8
Entity: Universitat Rovira i Virgili
Journal publication year: 2021
Publication Type: Journal Publications
Repositori URV