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    Direct electrochemical genosensing for multiple point mutation detection of Mycobacterium tuberculosis during the development of rifampin resistance
    (Elsevier Advanced Technology, 2009) Kara, Pinar; Cavusoglu, Cengiz; Cavdar, Seda; Ozsoz, Mehmet
    We present a robust and simple method for the direct detection of multiple point mutations in the Mycobacterium tuberculosis rpoB gene during the development of rifampin (RIF) resistance using an electrochemical genosensor. The device contained five different capture probes which are designed to hybridize with several sequence segments within the bacterial rpoB gene hotspot region. Point mutations were detected by monitoring the guanine oxidation with differential pulse voltammetry after hybridization between PCR amplicons and inosine modified capture probes at graphite surface. Changes in the peak voltage corresponding to guanine oxidation provide an electrochemical signal for hybridization that can be used to determine the presence of point mutations conferring rifampin resistance. The analytical parameters (sensitivity, selectivity and reproducibility) were evaluated. High selective discrimination against point mutation of bacteria at hot-spot region was observed. Several mutations were detected at several parts of the amplicon from 21 positive samples. (C) 2008 Elsevier B.V. All rights reserved.
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    Electrochemical genoassay design for allele-specific detection of toll-like receptor-2 gene polymorphism
    (Wiley-V C H Verlag Gmbh, 2007) Kara, Pinar; Cavdar, Seda; Berdeli, Afig; Ozsoz, Mehmet
    An allele-specific voltammetric genoassay for the detection of allele-specific toll-like receptor-2 gene arg753gln polymorphism (TLR-2) from polymerase chain reaction (PCR) amplified real samples was described in this study. Meldola blue (MDB), an intercalator molecule, was used as hybridization label. The wild-type and mutant type oligonucleotide probes were immobilized onto disposable graphite electrode surfaces by covalent attachment method. The extent of hybridization between probe and target sequences was determined by using differential pulse voltammetry (DPV). As a result of the interaction between MDB and DNA at electrode surface, the MDB signal observed from probe sequence before hybridization and after hybridization with MM sequence is lower than that observed after hybridization with complementary sequence. The differences between the MDB reduction peaks obtained from probe modified, hybrid modified and MM modified electrode were used to detect TLR-2 from PCR amplified real samples. The discrimination of homozygous and heterozygous alleles was also established by comparing the peak currents of MDB reduction signals. Numerous factors affecting the target hybridization and indicator binding reactions are optimized to maximize the sensitivity.
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    Electrochemical probe DNA design in PCR amplicon sequence for the optimum detection of microbiological diseases
    (Elsevier Science Sa, 2007) Kara, Pinar; Cavdar, Seda; Meric, Buket; Erensoy, Selda; Ozsoz, Mehmet
    Direct electrochemical genosensor was developed for the detection of a probe sequence relative position in a PCR amplicon for the optimum detection of bacterial and microbiological diseases, in this study. The genosensor relies on a label-free electrochemical detection. The amino-linked inosine modified (guanine-free) coequal capture probes which were chosen from different parts of a PCR amplicon, immobilized on to disposable pencil graphite electrodes (PGE) by electrostatically and covalently. As a model case Hepatitis B virus (HBV) genome amplicon was used for the detection and specification. Hybridization was occurred after surface coverage with denatured amplicons. After hybridization, optimum probe sequence position was identified by using the differences between the responses of guanine oxidation signals. The results of this study might have a great convenience for the microbiological diseases detection applications such as DNA micro arrays. (c) 2007 Elsevier B.V. All rights reserved.