Testing dough properties has been an essential part of quality evaluation of variety registration trials for Canada Western Red Spring (CWRS) wheat class. The balance between the elasticity and extensibility of dough, controlled by the composition of gluten proteins, determines processing properties and end-product quality. To achieve the desired balance, dough characteristics should be neither overly elastic nor weak. Carberry and Glenn are current check varieties used to reflect the floor and ceiling, respectively, of dough strength within which new lines should be targeted. In the 2017 and 2018 Western Bread Wheat Cooperative trials, check varieties and lines were grown at 13 locations across Western Canada. A small quantity of wheat of each check variety from each location was graded to determine an overall composite formulation for subsequent large scale quality evaluation. A small-scale rapid quality testing protocol, recently developed in our laboratory, was used to test these limited quantities of wheat from each location to investigate the impact of environment on dough strength. In order to elucidate the biochemical basis of the impact of environment, flour proteins were fractionated and quantified for total monomeric proteins, total glutenin, soluble glutenin and insoluble glutenin. The ratio of high molecular weight (HMW-) to low molecular weight (LMW-) glutenin subunits (GS), and the relative proportions of individual HMW-GS were analyzed with reversed phase-ultra high performance liquid chromatography. Results showed that the range of strength between the two check varieties changed with location, and in some locations there was no significant difference or even a switch in ranking between the floor and ceiling checks. These results confirm that strength is a result of genetics, environment and GxE interactions, and the importance of making composites from multiple locations for quality evaluation to ensure representative ranges and values for strength. For the sites with normal ranking of strength between the two checks, the amount of total monomeric proteins and insoluble glutenin in Glenn were consistently lower and higher, respectively, than those of Carberry. For locations with little difference or a switch in strength, the distribution of these protein fractions shifted in favor of increasing the strength of Carberry. The ratio of HMW- to LMW-GS and the relative proportion of Glu-1Dx subunit provided further insight into the variation in gluten strength between the two checks grown in various locations.