Katja M. Hoedjes (Amsterdam Institute for Life and Environment Vrije Universiteit Amsterdam Amsterdam The Netherlands) Katja M. Hoedjes Michael G. Ritchie (Centre for Biological Diversity University of St Andrews St Andrews UK) Michael G. Ritchie Christian Schlötterer Christian Schlötterer Jessica K. Abbott (Biology Department Lund University Lund Sweden) Jessica K. Abbott Isabel Almudi (Departament de Genètica, Microbiologia i Estadística Universitat de Barcelona Barcelona Spain) Isabel Almudi Marta Coronado‐Zamora (Institute of Evolutionary Biology CSIC, UPF Barcelona Spain) Marta Coronado‐Zamora Esra Durmaz Mitchell (Department of Biology University of Fribourg Fribourg Switzerland) Esra Durmaz Mitchell Thomas Flatt (Department of Biology University of Fribourg Fribourg Switzerland) Thomas Flatt Claudia Fricke (Institute for Zoology/Animal Ecology Martin‐Luther‐University Halle‐Wittenberg Halle (Saale) Germany) Claudia Fricke Amanda Glaser‐Schmitt (Division of Evolutionary Biology LMU Munich Planegg‐Martinsried Germany) Amanda Glaser‐Schmitt Josefa González (Institute of Evolutionary Biology CSIC, UPF Barcelona Spain) Josefa González Luke Holman (School of Applied Sciences Edinburgh Napier University Edinburgh UK) Luke Holman Maaria Kankare (Department of Biological and Environmental Science University of Jyväskylä Jyväskylä Finland) Maaria Kankare Benedict Lenhart (Department of Biology University of Virginia Charlottesville Virginia USA) Benedict Lenhart Dorcas J. Orengo (Departament de Genètica, Microbiologia i Estadística Universitat de Barcelona Barcelona Spain) Dorcas J. Orengo Rhonda R. Snook (Department of Zoology Stockholm University Stockholm Sweden) Rhonda R. Snook Vera M. Yılmaz (Division of Evolutionary Biology LMU Munich Planegg‐Martinsried Germany) Vera M. Yılmaz Leeban Yusuf (Centre for Biological Diversity University of St Andrews St Andrews UK) Leeban Yusuf Sonja Grath Sonja Grath Nico Posnien Nico Posnien Wiley 2025 RNA sequencing (RNAseq) methodology has experienced a burst of technological developments in the last decade, which has opened up opportunities for studying the mechanisms of adaptation to environmental factors at both the organismal and cellular level. Selecting the most suitable experimental approach for specific research questions and model systems can, however, be a challenge and researchers in ecology and evolution are commonly faced with the choice of whether to study gene expression variation in whole bodies, specific tissues, and/or single cells. A wide range of sometimes polarised opinions exists over which approach is best. Here, we highlight the advantages and disadvantages of each of these approaches to provide a guide to help researchers make informed decisions and maximise the power of their study. Using illustrative examples of various ecological and evolutionary research questions, we guide the readers through the different RNAseq approaches and help them identify the most suitable design for their own projects. PDFDocument eng 2026-01-30T14:44:30.922737Z 2025 Bulk RNAseq Cellular Heterogeneity Deconvolution Gene Expression Single-cell RNAseq Transcriptomics 1109573 b application/pdf http://creativecommons.org/licenses/by/4.0/