Abstract
Plant-derived extracellular vesicles (EVs), containing a myriad of bioactive proteins, microRNAs, lipids, and secondary metabolites, have recently become the focus of rising interest due to their important roles in various applications. The widely accepted method for isolating plant EVs is differential ultracentrifugation plus density gradient centrifugation. However, the combination of differential ultracentrifugation and density gradient centrifugation for the isolation of plant EVs is time-consuming and labor-intensive. Hence, there is a need for more efficient methods to perform the separation of plant EVs. In this study, EVs were separated from Arabidopsis thaliana leaves by a cost-effective polyethylene glycol (PEG)-based precipitation approach. The mean size of purified Arabidopsis thaliana EVs determined by dynamic light scattering was 266 nm, which is consistent with nanoparticle tracking analysis. The size was also confirmed via transmission electron microscopy with morphology of a cup-shaped appearance which is the typical mammalian exosome’s morphology. Additionally, Western blotting of the purified Arabidopsis thaliana EVs, using commercially available mammalian exosomal kits, displayed surface marker tetraspanin proteins (CD9, CD63, and CD81), and endosomal sorting complexes required for transport (ESCRT)-associated proteins (TSG101 and ALIX). This demonstrates that the purified Arabidopsis thaliana EVs reveal the typical proteins reported in mammalian exosomes.
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The data that support the findings of this study are available from the corresponding author upon reasonable request.
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This research is supported by the National Institute of Food and Agriculture – AFRI Project 1028230 and Hatch Project 7007183.
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Sharjeel Jokhio: methodology, investigation, formal analysis, validation, and writing—original draft. Ian Peng: methodology, investigation, formal analysis, validation, and writing—original draft. Ching-An Peng: conceptualization, methodology, investigation, formal analysis, validation, supervision, funding acquisition, project administration, resources, and writing—review and editing.
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Communicated by Handling Editor: Liwen Jiang.
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Jokhio, S., Peng, I. & Peng, CA. Extracellular vesicles isolated from Arabidopsis thaliana leaves reveal characteristics of mammalian exosomes. Protoplasma (2024). https://doi.org/10.1007/s00709-024-01954-x
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DOI: https://doi.org/10.1007/s00709-024-01954-x