Background: Inclusion of racial/ethnic minorities in cancer-related research remains inadequate, continuing to impact disparate health outcomes. Data regarding enrollment of racial/ethnic minorities in nontherapeutic, health-related research is sparse, and even less is known about enrollment of those with a double disparity of racial/ethnic minorities in rural settings. Objective: To examine perspectives of Latinx and African American (AA) healthy volunteers from rural and urban settings in five southern US states and Puerto Rico regarding their trust in research and their willingness to participate in nontherapeutic research trials based on who conducts the research. Methods: This study was conducted in 2013 in rural and urban communities across Alabama, Florida, Georgia, Mississippi, Louisiana, and Puerto Rico. A 38-item questionnaire based on the Tuskegee Legacy Project Questionnaire assessed willingness, motivators, and barriers to participation in nontherapeutic cancer-related research. The sample was segmented into four subgroups by urban/rural location and race/ethnicity. Results: Of 553 participants (rural Latinx=151, urban Latinx=158, rural AA=122, urban AA=122), more than 90% had never been asked to participate in research, yet 75% of those asked agreed to participate. Most had positive views regarding health-related research. Trust in who conducted research varied by subgroup. Personal doctors and university hospitals were most trusted by all subgroups; for-profit and tobacco companies were least trusted. Both Latinx subgroups trusted pharmaceutical companies more than AAs; local hospitals and for-profit businesses were more trusted by AAs. Both rural subgroups trusted research by insurance companies more than their urban counterparts. Conclusions: If asked, rural and urban AA and Latinx healthy volunteers were willing to participate in health-related research, with personal doctors and university hospitals considered the most trusted sources to encourage/conduct research.