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Alfalfa (Medicago sativa L.) was the first perennial crop genetically engineered (GE) to resist glyphosate herbicide. Since alfalfa is insect pollinated, the potential for transgene flow and occurrence of adventitious presence (AP) in conventional varieties is a concern, especially for organic and export producers. In response, the industry developed seed production isolation distances. However, no landscape‐scale studies have been conducted to validate industry standards. Our objective was to quantify the extent of gene movement from GE seed fields to commercial conventional seed fields on a landscape level to provide the industry with information to support coexistence strategies. Alfalfa seed fields were mapped in the Touchet area, Walla Walla Valley, Washington, during spring 2013. Fourteen conventional seed fields located at various distances from GE sources were identified, harvested, and tested for the presence of the GE trait. Although other variables affect gene flow, this article focuses on geographic distance, since this variable can be easily controlled by growers and industry. Using distance as the predictor variable, we modeled the relationship between AP proportion and distance to GE source fields at 0.1, 0.5, and 0.9% AP threshold levels using quadratic regression. As distance increased, transgene flow generally decreased. At a threshold level of 0.9% AP, an isolation distance of 330 m between conventional and GE fields is needed to ensure that at least 95% of the samples fall below this threshold level. Spatial prediction maps were consistent with our statistical analysis. At an average distance of 2.2 km, AP was 0.05%, and this decreased to 0.03% at an average distance of 8.8 km from GE seed fields. [ABSTRACT FROM AUTHOR]