Molecular Ecology open access doi:10.1111/mec.17382 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. Text Wissenschaftlicher Artikel Wiley http://creativecommons.org/licenses/by/4.0/ © 2024 The Author(s) application/pdf eng Testo Articolo di rivista Text journal article Bulk RNAseq Cellular Heterogeneity Deconvolution Gene Expression Single-cell RNAseq Transcriptomics Katja M. Hoedjes (Amsterdam Institute for Life and Environment Vrije Universiteit Amsterdam Amsterdam The Netherlands) Michael G. Ritchie (Centre for Biological Diversity University of St Andrews St Andrews UK) Christian Schlötterer Jessica K. Abbott (Biology Department Lund University Lund Sweden) Isabel Almudi (Departament de Genètica, Microbiologia i Estadística Universitat de Barcelona Barcelona Spain) Marta Coronado‐Zamora (Institute of Evolutionary Biology CSIC, UPF Barcelona Spain) Esra Durmaz Mitchell (Department of Biology University of Fribourg Fribourg Switzerland) Thomas Flatt (Department of Biology University of Fribourg Fribourg Switzerland) Claudia Fricke (Institute for Zoology/Animal Ecology Martin‐Luther‐University Halle‐Wittenberg Halle (Saale) Germany) Amanda Glaser‐Schmitt (Division of Evolutionary Biology LMU Munich Planegg‐Martinsried Germany) Josefa González (Institute of Evolutionary Biology CSIC, UPF Barcelona Spain) Luke Holman (School of Applied Sciences Edinburgh Napier University Edinburgh UK) Maaria Kankare (Department of Biological and Environmental Science University of Jyväskylä Jyväskylä Finland) Benedict Lenhart (Department of Biology University of Virginia Charlottesville Virginia USA) Dorcas J. Orengo (Departament de Genètica, Microbiologia i Estadística Universitat de Barcelona Barcelona Spain) Rhonda R. Snook (Department of Zoology Stockholm University Stockholm Sweden) Vera M. Yılmaz (Division of Evolutionary Biology LMU Munich Planegg‐Martinsried Germany) Leeban Yusuf (Centre for Biological Diversity University of St Andrews St Andrews UK) Sonja Grath Nico Posnien From whole bodies to single cells: A guide to transcriptomic approaches for ecology and evolutionary biology https://phaidra.vetmeduni.ac.at/o:4836