Details zur Publikation
Kategorie Textpublikation
Referenztyp Buchkapitel
DOI 10.1007/978-1-4939-7634-8_13
Titel (primär) Extraction and analysis of RNA isolated from pure bacteria-derived outer membrane vesicle
Titel (sekundär) Bacterial regulatory RNA : methods and protocols
Autor Habier, J.; May, P.; Heintz-Buschart, A.; Ghosal, A.; Wienecke-Baldacchino, A.K.; Nolte-‘t Hoen, E.N.M.; Wilmes, P.; Fritz, J.V.
Herausgeber Arluison, V.; Valverde, C.
Quelle Methods in Molecular Biology
Erscheinungsjahr 2018
Department BOOEK; iDiv
Band/Volume 1737
Seite von 213
Seite bis 230
Sprache englisch
Keywords Bacteria; RNA; Outer membrane vesicle (OMV); Gram-negative; Sequencing; Analysis; Extraction; Ultracentrifugation; Ultrafiltration; Density gradient
Abstract Outer membrane vesicles (OMVs) are released by commensal as well as pathogenic Gram-negative bacteria. These vesicles contain numerous bacterial components, such as proteins, peptidoglycans, lipopolysaccharides, DNA, and RNA. To examine if OMV-associated RNA molecules are bacterial degradation products and/or are functionally active, it is necessary to extract RNA from pure OMVs for subsequent analysis. Therefore, we describe here an isolation method of ultrapure OMVs and the subsequent extraction of RNA and basic steps of RNA-Seq analysis. Bacterial culture, extracellular supernatant concentration, OMV purification, and the subsequent RNA extraction out of OMVs are described. Specific pitfalls within the protocol and RNA contamination sources are highlighted.
dauerhafte UFZ-Verlinkung https://www.ufz.de/index.php?en=20939&ufzPublicationIdentifier=21384
Habier, J., May, P., Heintz-Buschart, A., Ghosal, A., Wienecke-Baldacchino, A.K., Nolte-‘t Hoen, E.N.M., Wilmes, P., Fritz, J.V. (2018):
Extraction and analysis of RNA isolated from pure bacteria-derived outer membrane vesicle
In: Arluison, V., Valverde, C. (eds.)
Bacterial regulatory RNA : methods and protocols
Methods in Molecular Biology 1737
Springer Nature, p. 213 - 230 10.1007/978-1-4939-7634-8_13