Standard operating procedure for next generation sequencing of RNA isolated form extracellular vesicles within the nanostructurome methods pipeline

Kovachevikj, Miona (author), Kralj-Iglič, Veronika (author), Jakše, Jernej (author), Jeseničnik, Taja (author)

The standard operating procedure (SOP) for next generation sequencing (NGS) of RNA from extracellular vesicles is presented. The SOP enables purification of small or total RNAs from isolated extracellular vesicles. Next, the SOP describes the sequencing of RNAs using NGS sequencing platform IonTorrent. In the presented protocol we define all the steps from isolation of RNA to sequencing data management, together with the cautions and checkpoints crucial for obtaining reliable high-quality sequencing reads, suitable for further analysis. NGS enables high-throughput DNA and RNA sequencing, offering accuracy, scalability, and cost-efficiency. RNA sequencing is widely used for analysing gene expression, alternative splicing, and non-coding RNAs. NGS also facilitates the study of extracellular vesicles (EVs), which play a key role in intercellular communication, especially in plant-pathogen interactions. Insights gained could inform novel disease management strategies, such as Spray-Induced Gene Silencing, for improved crop protection.

• Language: English
• Keywords: next generation sequencing, extracellular vesicles, RNA-seq, total RNA, small RNA, IonTorrent
• Work type: Scientific work
• Typology: 1.08 - Published Scientific Conference Contribution
• Organization: ZF - Faculty of Health Sciences; BF - Biotechnical Faculty
• Publication status: Published
• Publication version: Version of Record
• Year: 2025
• Number of pages: Str. 18-24
• PID: 20.500.12556/RUL-169993
• UDC: 577
• DOI: 10.55295/PSL.12.2025.III3
• COBISS.SI-ID: 240973827
• Publication date in RUL: 01.07.2025

Record is a part of a monograph

• Title: Socratic Lectures : 12th international symposium, Ljubljana, 11. 1. 2025
• Editors: Veronika Kralj-Iglič, Yelena Istileulova, Anna Romolo
• Place of publishing: Ljubljana
• Publisher: University of Ljubljana Press
• Year: 2025
• ISBN: 978-961-297-597-5
• COBISS.SI-ID: 237747715

Licences

• License: CC BY 4.0, Creative Commons Attribution 4.0 International
• Link: http://creativecommons.org/licenses/by/4.0/
• Description: This is the standard Creative Commons license that gives others maximum freedom to do what they want with the work as long as they credit the author.

Projects

• Funder: ARIS - Slovenian Research and Innovation Agency
• Project number: P4-0077-2019
• Name: Kmetijske rastline - genetika in sodobne tehnologije

• Funder: ARIS - Slovenian Research and Innovation Agency
• Project number: 802-12/2024-5, J3-60063,
• Name: Nanostructurome