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      Mismatch detection in DNA monolayers by atomic force microscopy and electrochemical impedance spectroscopy

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          Summary

          Background: DNA hybridization is at the basis of most current technologies for genotyping and sequencing, due to the unique properties of DNA base-pairing that guarantee a high grade of selectivity. Nonetheless the presence of single base mismatches or not perfectly matched sequences can affect the response of the devices and the major challenge is, nowadays, to distinguish a mismatch of a single base and, at the same time, unequivocally differentiate devices read-out of fully and partially matching sequences.

          Results: We present here two platforms based on different sensing strategies, to detect mismatched and/or perfectly matched complementary DNA strands hybridization into ssDNA oligonucleotide monolayers. The first platform exploits atomic force microscopy-based nanolithography to create ssDNA nano-arrays on gold surfaces. AFM topography measurements then monitor the variation of height of the nanostructures upon biorecognition and then follow annealing at different temperatures. This strategy allowed us to clearly detect the presence of mismatches. The second strategy exploits the change in capacitance at the interface between an ssDNA-functionalized gold electrode and the solution due to the hybridization process in a miniaturized electrochemical cell. Through electrochemical impedance spectroscopy measurements on extended ssDNA self-assembled monolayers we followed in real-time the variation of capacitance, being able to distinguish, through the difference in hybridization kinetics, not only the presence of single, double or triple mismatches in the complementary sequence, but also the position of the mismatched base pair with respect to the electrode surface.

          Conclusion: We demonstrate here two platforms based on different sensing strategies as sensitive and selective tools to discriminate mismatches. Our assays are ready for parallelization and can be used in the detection and quantification of single nucleotide mismatches in microRNAs or in genomic DNA.

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          Ionic liquids at electrified interfaces.

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            DNA microarray technology: devices, systems, and applications.

            In this review, recent advances in DNA microarray technology and their applications are examined. The many varieties of DNA microarray or DNA chip devices and systems are described along with their methods for fabrication and their use. This includes both high-density microarrays for high-throughput screening applications and lower-density microarrays for various diagnostic applications. The methods for microarray fabrication that are reviewed include various inkjet and microjet deposition or spotting technologies and processes, in situ or on-chip photolithographic oligonucleotide synthesis processes, and electronic DNA probe addressing processes. The DNA microarray hybridization applications reviewed include the important areas of gene expression analysis and genotyping for point mutations, single nucleotide polymorphisms (SNPs), and short tandem repeats (STRs). In addition to the many molecular biological and genomic research uses, this review covers applications of microarray devices and systems for pharmacogenomic research and drug discovery, infectious and genetic disease and cancer diagnostics, and forensic and genetic identification purposes. Additionally, microarray technology being developed and applied to new areas of proteomic and cellular analysis are reviewed.
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              The common colorectal cancer predisposition SNP rs6983267 at chromosome 8q24 confers potential to enhanced Wnt signaling.

              Homozygosity for the G allele of rs6983267 at 8q24 increases colorectal cancer (CRC) risk approximately 1.5 fold. We report here that the risk allele G shows copy number increase during CRC development. Our computer algorithm, Enhancer Element Locator (EEL), identified an enhancer element that contains rs6983267. The element drove expression of a reporter gene in a pattern that is consistent with regulation by the key CRC pathway Wnt. rs6983267 affects a binding site for the Wnt-regulated transcription factor TCF4, with the risk allele G showing stronger binding in vitro and in vivo. Genome-wide ChIP assay revealed the element as the strongest TCF4 binding site within 1 Mb of MYC. An unambiguous correlation between rs6983267 genotype and MYC expression was not detected, and additional work is required to scrutinize all possible targets of the enhancer. Our work provides evidence that the common CRC predisposition associated with 8q24 arises from enhanced responsiveness to Wnt signaling.
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                Author and article information

                Contributors
                Role: Guest Editor
                Journal
                Beilstein J Nanotechnol
                Beilstein J Nanotechnol
                Beilstein Journal of Nanotechnology
                Beilstein-Institut (Trakehner Str. 7-9, 60487 Frankfurt am Main, Germany )
                2190-4286
                2016
                9 February 2016
                : 7
                : 220-227
                Affiliations
                [1 ]Elettra-Sincrotrone Trieste S.C.p.A., s.s. 14 km 163.5 in Area Science Park, Basovizza, Trieste, Italy
                [2 ]INSTM – ST Unit, s.s. 14 km 163.5 in Area Science Park, Basovizza, Trieste, Italy
                [3 ]University of Trieste, Via Valerio 9, Trieste, Italy
                [4 ]Department of Medical and Biological Sciences, University of Udine, Udine, Italy
                Article
                10.3762/bjnano.7.20
                4778512
                26977379
                ec913cdb-0cf3-4a8f-828f-878c5954d2c3
                Copyright © 2016, Ngavouka et al; licensee Beilstein-Institut.

                This is an Open Access article under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

                The license is subject to the Beilstein Journal of Nanotechnology terms and conditions: ( http://www.beilstein-journals.org/bjnano)

                History
                : 19 October 2015
                : 25 January 2016
                Categories
                Full Research Paper
                Nanoscience
                Nanotechnology

                atomic force microscopy,dna monolayers,electrochemical impedance spectroscopy,hybridization,mismatches

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