End-point RT-PCR based on a conservation landscape for SARS-COV-2 detection

Autor: Armando Cruz-Rangel, Laura Gómez-Romero, Mireya Cisneros-Villanueva, G. de Anda Jáuregui, Victor Luna-Pineda, Alberto Cedro-Tanda, Abraham Campos-Romero, Alfredo Mendoza-Vargas, J. P. Reyes-Grajeda, Alfredo Hidalgo-Miranda, COVID-19 Consortium INMEGEN, Luis A. Herrera, Felipe Vadillo-Ortega
Jazyk: angličtina
Rok vydání: 2022
Předmět:
Zdroj: Scientific Reports, Vol 12, Iss 1, Pp 1-13 (2022)
Druh dokumentu: article
ISSN: 2045-2322
DOI: 10.1038/s41598-022-07756-6
Popis: Abstract End-point RT-PCR is a suitable alternative diagnostic technique since it is cheaper than RT-qPCR tests and can be implemented on a massive scale in low- and middle-income countries. In this work, a bioinformatic approach to guide the design of PCR primers was developed, and an alternative diagnostic test based on end-point PCR was designed. End-point PCR primers were designed through conservation analysis based on kmer frequency in SARS-CoV-2 and human respiratory pathogen genomes. Highly conserved regions were identified for primer design, and the resulting PCR primers were used to amplify 871 nasopharyngeal human samples with a previous RT-qPCR based SARS-CoV-2 diagnosis. The diagnostic test showed high accuracy in identifying SARS-CoV-2-positive samples including B.1.1.7, P.1, B.1.427/B.1.429 and B.1.617.2/ AY samples with a detection limit of 7.2 viral copies/µL. In addition, this test could discern SARS-CoV-2 infection from other viral infections with COVID-19-like symptomatology. The designed end-point PCR diagnostic test to detect SARS-CoV-2 is a suitable alternative to RT-qPCR. Since the proposed bioinformatic approach can be easily applied in thousands of viral genomes and over highly divergent strains, it can be used as a PCR design tool as new SARS-CoV-2 variants emerge. Therefore, this end-point PCR test could be employed in epidemiological surveillance to detect new SARS-CoV-2 variants as they emerge and propagate.
Databáze: Directory of Open Access Journals
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