Recent research suggests that underrepresented minority (URM) college students, and especially first-generation URMs, may lose motivation to persist if they see science careers as unable to fulfill culturally relevant career goals. In the present study, we used a mixed-methods approach to explore patterns of motivation to pursue physical and life sciences across ethnic groups of freshman college students, as moderated by generational status. Results from a longitudinal survey (N = 249) demonstrated that freshman URM students who enter with a greater belief that science can be used to help their communities identified as scientists more strongly over time, but only among first-generation college students. Analysis of the survey data were consistent with content analysis of 11 transcripts from simultaneously conducted focus groups (N = 67); together, these studies reveal important differences in motivational characteristics both across and within ethnicity across educational generation status. First-generation URM students held the strongest prosocial values for pursuing a science major (e.g., giving back to the community). URM students broadly reported additional motivation to increase the status of their family (e.g., fulfilling aspirations for a better life). These findings demonstrate the importance of culturally connected career motives and for examining intersectional identities to understand science education choices and inform efforts to broaden participation.