Effect of fieldwork-friendly coffee blender-based extraction methods and leaf tissue storage on the transcriptome of non-model plants

TL;DR

This study evaluates field-friendly methods for RNA extraction and storage in plants, focusing on their impact on transcriptome quality in non-model species.

Contribution

The study introduces and validates fieldwork-friendly RNA extraction protocols suitable for polyphenol-rich plants and non-cold storage conditions.

Findings

Transcriptome assemblies from stored leaf tissue and field extraction methods showed quality comparable to state-of-the-art methods.

Onsite RNA extraction methods outperformed stored tissue samples in differential expression analysis.

Protocols were optimized for use in remote areas without cold storage or advanced equipment.

Abstract

The adaptation of plants to environmental conditions involves a transcriptional response. “Field transcriptomics” is an emerging concept for studying plants in their natural habitat. However, this term includes studies in which cold storage was possible until further processing in a laboratory. Previous studies proposing onsite RNA extraction methods are limited to descriptions of RNA purity, quantity, and quality, and lack a thorough evaluation of transcriptome quality, and transcriptomic evaluations of RNA storage solutions in plants are, to our knowledge, only available for periods of less than a day. This issue is critical for studying plants in geographically difficult-to-access regions, where keeping the cold chain is unrealistic. In this study, the transcriptome of the non-model plant Helonias orientalis (order: Liliales) was evaluated before and after storage of the leaf tissue for one and fourteen days at 25 °C in RNAlater and TRIzol, respectively. Additionally, field-friendly protocols were similarly evaluated for onsite plant RNA extraction at ambient temperature with lightweight equipment that can run on a portable generator, including a guanidine isothiocyanate-free protocol that is compatible with the polyphenol-rich wild strawberry Fragaria vesca. The quality of the transcriptome assembly after 1-day storage and our optimized onsite methods had similar results to that of the state-of-the-art. However, in terms of differential expression analysis, onsite extraction methods performed better overall than the stored tissue samples. We expect that our onsite RNA extraction methods will provide valuable insights into the transcriptional regulation of plants in areas where research equipment is difficult to access.

The online version contains supplementary material available at 10.1007/s10265-025-01624-w.

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