Evaluation of TaqMan qPCR system integrating two identically labelled hydrolysis probes in single assay

Result code in IS VaVaI

<a href="https://www.isvavai.cz/riv?ss=detail&h=RIV%2F75010330%3A_____%2F17%3A00011792" target="_blank" >RIV/75010330:_____/17:00011792 - isvavai.cz</a>

Result on the web

<a href="http://www.nature.com/articles/srep41392" target="_blank" >http://www.nature.com/articles/srep41392</a>

DOI - Digital Object Identifier

<a href="http://dx.doi.org/10.1038/srep41392" target="_blank" >10.1038/srep41392</a>

Result language

angličtina

Original language name

Evaluation of TaqMan qPCR system integrating two identically labelled hydrolysis probes in single assay

Original language description

Ongoing evolution of viral pathogens is a significant issue in diagnostic virology employing TaqMan qPCR/RT-qPCR. Specific concerns are related to false negativity due to probe binding failure. One option for compensating for such deficiency is to integrate a second identically labelled probe in the assay. However, how this alteration influences the reaction parameters has not been comprehensively demonstrated. In the present study, we evaluate a TaqMan protocol using two identically labelled hydrolysis probes (simple, LNA (locked-nucleic-acid)) and MGB (minor-groove-binder) modified probes and combinations thereof in a single assay. Our results based on a synthetic amplicon suggest that the second probe does not compromise the TaqMan qPCR/RT-qPCR parameters, which repeatedly and reproducibly remained comparable to those of the corresponding single-probe assays, irrespective of the relative probe orientation, whether opposite or tandem, and probe modifications or combinations thereof. On the other hand, the second probe additively contributed to the overall fluorescence signal. The utility of the dual-probe approach was demonstrated on practical examples by using field specimens. We hope that the present study might serve as a theoretical basis for the development or improvement of TaqMan qPCR/RT-qPCR assays for the detection of highly variable nucleic acid templates.

Czech name

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Czech description

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Type

J_imp - Article in a specialist periodical, which is included in the Web of Science database

CEP classification

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OECD FORD branch

30303 - Infectious Diseases

Project

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Continuities

I - Institucionalni podpora na dlouhodoby koncepcni rozvoj vyzkumne organizace

Publication year

2017

Confidentiality

S - Úplné a pravdivé údaje o projektu nepodléhají ochraně podle zvláštních právních předpisů

Name of the periodical

Scientific Reports

ISSN

2045-2322

e-ISSN

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Volume of the periodical

7

Issue of the periodical within the volume

January

Country of publishing house

GB - UNITED KINGDOM

Number of pages

10

Pages from-to

nestrankovano

UT code for WoS article

000392592300001

EID of the result in the Scopus database

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