© 2021, The Author(s), under exclusive licence to Springer Nature B.V.The purpose of the study was to examine science teachers’ emotions, emotion regulation goals and strategies during instruction, and the role of teaching experience, teacher efficacy beliefs, and teacher goal orientations in their emotions and emotion regulation using the control-value theory of emotions and the process model of emotion regulation. We employed a multiple-case holistic design with three in-service science teachers. Data were collected from teacher diaries, video recordings, fieldnotes from non-participant observations, and semi-structured interviews. For data analysis, the deductive method, content analysis, and the constant comparative method were employed. Results showed that the most frequently experienced emotions during the instruction were enjoyment and contentment as positive emotions and worry and anger as negative emotions. The teachers mostly used antecedent-focused emotion regulation strategies for hedonic reasons. The most frequently used emotion regulation strategy was situation selection, which was mostly employed for classroom management issues. In addition, findings indicated that the teachers differed in their emotions, emotion regulation goals, and emotion regulation strategies based on their years of teaching experience, self-efficacy beliefs, goal orientations, and pedagogical content knowledge. Results suggest the importance of teacher emotions in science education to nurture effective science pedagogies and to enrich science teaching.