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http://hdl.handle.net/2445/207048
Title: | Comparison of Extracellular Vesicle Isolation Methods for miRNA Sequencing |
Author: | Llorens-Revull, Meritxell Martínez-González, Brenda Quer, Josep Esteban, Juan Ignacio Núñez-Moreno, Gonzalo Mínguez, Pablo Burgui, Idoia Ramos-Ruíz, Ricardo Soria, María Eugenia Rico, Angie Riveiro-Barciela, Mar Sauleda, Silvia Piron, María Corrales, Irene Borràs i Serres, Francesc Enric Rodríguez-Frías, Francisco Rando, Ariadna Ramírez-Serra, Clara Camós, Silvia Domingo, Esteban Bes, Marta Perales, Celia Costafreda Salvany, M. Isabel (Maria Isabel) |
Keywords: | RNA Marcadors bioquímics RNA Biochemical markers |
Issue Date: | 29-Jul-2023 |
Publisher: | MDPI |
Abstract: | MicroRNAs (miRNAs) encapsulated in extracellular vesicles (EVs) are potential diagnostic and prognostic biomarkers. However, discrepancies in miRNA patterns and their validation are still frequent due to differences in sample origin, EV isolation, and miRNA sequencing methods. The aim of the present study is to find a reliable EV isolation method for miRNA sequencing, adequate for clinical application. To this aim, two comparative studies were performed in parallel with the same human plasma sample: (i) isolation and characterization of EVs obtained using three procedures: size exclusion chromatography (SEC), iodixanol gradient (GRAD), and its combination (SEC+GRAD) and (ii) evaluation of the yield of miRNA sequences obtained using NextSeq 500 (Illumina) and three miRNA library preparation protocols: NEBNext, NEXTFlex, and SMARTer smRNA-seq. The conclusion of comparison (i) is that recovery of the largest amount of EVs and reproducibility were attained with SEC, but GRAD and SEC+GRAD yielded purer EV preparations. The conclusion of (ii) is that the NEBNext library showed the highest reproducibility in the number of miRNAs recovered and the highest diversity of miRNAs. These results render the combination of GRAD EV isolation and NEBNext library preparation for miRNA retrieval as adequate for clinical applications using plasma samples. |
Note: | Reproducció del document publicat a: https://doi.org/10.3390/ijms241512183 |
It is part of: | International Journal of Molecular Sciences, 2023, vol. 24, num.15, p. 1-15 |
URI: | http://hdl.handle.net/2445/207048 |
Related resource: | https://doi.org/10.3390/ijms241512183 |
ISSN: | 1661-6596 |
Appears in Collections: | Articles publicats en revistes (Genètica, Microbiologia i Estadística) |
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