This data set documents the gene transcription levels for a panel of 12 selected genes and the heat shock protein 70 (HSP70) protein abundance measured in the muscle tissue of individual wild Chinook salmon captured from locations within the U.S. portion of the Yukon River watershed. Chinook salmon were primarily captured in 2016 and 2017 from existing field efforts (n = 477). A small number of additional samples (n = 22) were collected in 2018 following an experimental temperature experiment conducted in Pilot Station, AK, USA. During this experiment individual fish were held for ~6 hours at either river ambient control water temperatures or a heated treatment temperature of 18 °C or 21 °C. This small experiment data set was used to validate and refine the use of gene transcription and HSP70 protein levels as heat stress biomarkers for Yukon River Chinook salmon. The analysis of this data was published in von Biela et al. 2020 (DOI:10.1139/cjfas-2020-0209).