Usefulness of an in vitro-transcribed RNA control for the detection and quantification of Yellow fever virus through real-time reverse transcription-polymerase chain reaction

Introduction: Unvaccinated individuals in endemic areas with proven enzootic transmission of Yellow fever virus are at risk of infection due to a dramatic shift in the epidemiology of the disease over recent years. For this reason, epidemiological surveillance and laboratory confirmation of cases have become mandatory. Objective: To develop and test a control RNA for YFV detection through real-time RT-PCR. Methods: A 437-bp insert containing the T7 promoter and the target sequences for two different in-house protocols was designed in the context of the pUC57 vector and obtained through gene synthesis. After T7-driven in vitro transcription, standard curves were developed for Log10 serial dilutions of the YFV control RNA with 8 replicates. Results: A dynamic range of quantification of 10 orders of magnitude was observed with a limit of detection of 6.3 GCE/µL (95% CI, 2.6 to 139.4 GCE/µL). Conclusion: The plasmid construct is available for YFV molecular test validation on clinical, entomological, and epizootic samples.

• Autores: Laiton Donato, Katherine; Quintero Cortés, Paula; Franco Salazar, Juan Pablo; Peláez Carvajal, Dioselina; Navas, Maria Cristina; Junglen, Sandra; Parra Henao, Gabriel; Usme Ciro, José Aldemar
• Tipo de recurso: Article of journal
• Fecha de publicación: 2023
• Institución: Universidad Cooperativa de Colombia
• Repositorio: Repositorio UCC
• Idioma: eng
• OAI Identifier: oai:repository.ucc.edu.co:20.500.12494/55530
• Acceso en línea: https://hdl.handle.net/20.500.12494/55530
• Palabra clave: Yellow fever virus; RT-qPCR; Molecular detection; in vitro transcription; Plasmid DNA
• Rights: openAccess
• License: http://creativecommons.org/licenses/by-nc-nd/4.0/