Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
The result's identifiers
Result code in IS VaVaI
<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F61388963%3A_____%2F21%3A00550081" target="_blank" >RIV/61388963:_____/21:00550081 - isvavai.cz</a>
Alternative codes found
RIV/00216208:11310/21:10441839
Result on the web
<a href="https://doi.org/10.1021/acssensors.1c01710" target="_blank" >https://doi.org/10.1021/acssensors.1c01710</a>
DOI - Digital Object Identifier
<a href="http://dx.doi.org/10.1021/acssensors.1c01710" target="_blank" >10.1021/acssensors.1c01710</a>
Alternative languages
Result language
angličtina
Original language name
Electrochemical Detection of Single-Nucleotide Polymorphism Associated with Rifampicin Resistance in Mycobacterium tuberculosis Using Solid-Phase Primer Elongation with Ferrocene-Linked Redox-Labeled Nucleotides
Original language description
Here, we report the electrochemical detection of single-point mutations using solid-phase isothermal primer elongation with redox-labeled oligonucleotides. A single-base mutation associated with resistance to rifampicin, an antibiotic commonly used for the treatment of Mycobacterium tuberculosis, was used as a model system to demonstrate a proof-of-concept of the approach. Four 5′-thiolated primers, designed to be complementary with the same fragment of the target sequence and differing only in the last base, addressing the polymorphic site, were self-assembled via chemisorption on individual gold electrodes of an array. Following hybridization with single-stranded DNA, Klenow (exo-) DNA polymerase-mediated primer extension with ferrocene-labeled 2′-deoxyribonucleoside triphosphates (dNFcTPs) was only observed to proceed at the electrode where there was full complementarity between the surface-tethered probe and the target DNA being interrogated. We tested all four ferrocenylethynyl-linked dNTPs and optimized the ratio of labeled/natural nucleotides to achieve maximum sensitivity. Following a 20 min hybridization step, Klenow (exo-) DNA polymerase-mediated primer elongation at 37 °C for 5 min was optimal for the enzymatic incorporation of a ferrocene-labeled nucleotide, achieving unequivocal electrochemical detection of a single-point mutation in 14 samples of genomic DNA extracted from Mycobacterium tuberculosis strains. The approach is rapid, cost-effective, facile, and can be extended to multiplexed electrochemical single-point mutation genotyping.
Czech name
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Czech description
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Classification
Type
J<sub>imp</sub> - Article in a specialist periodical, which is included in the Web of Science database
CEP classification
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OECD FORD branch
10401 - Organic chemistry
Result continuities
Project
<a href="/en/project/GX20-00885X" target="_blank" >GX20-00885X: Novel Functionalized (Bio)polymers Based on DNA Display of Small Molecules</a><br>
Continuities
I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace
Others
Publication year
2021
Confidentiality
S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů
Data specific for result type
Name of the periodical
ACS Sensors
ISSN
2379-3694
e-ISSN
2379-3694
Volume of the periodical
6
Issue of the periodical within the volume
12
Country of publishing house
US - UNITED STATES
Number of pages
10
Pages from-to
4398-4407
UT code for WoS article
000755681200015
EID of the result in the Scopus database
2-s2.0-85120352225