Experienced teachers possess a unique teaching knowledge comprised of an inter-related set of knowledge and beliefs that gives direction and justification to a teacher's actions. This study examined the expansion of two components of pedagogical content knowledge (PCK) of three in-service teachers in the course of a professional development program aimed at designing new teaching and learning materials suggested by the teachers themselves. The research presents an enlargement of previous PCK representations by focusing on a detailed representation of two main PCK domains: teaching and learning, including ten PCK components that emerged in the course of data analysis. This representation enabled revealing the unique PCK held by each teacher and to characterize the expansion of the two components of the participating teachers' PCK during the long-term professional development program. Retention of major parts of the expanded PCK a year after termination of the program implies that designing and implementing new teaching and learning materials based on the teachers' experiences, needs, and knowledge in a workshop format accompanied by biology and science education courses might provide a powerful means for PCK expansion. We recommend that designers of professional development programs be aware of the unique PCK held by each teacher in order to promote meaningful professional development of each teacher. Moreover, the PCK representations that were identified in the course of this study enabled clarifying the "orientation toward teaching science" category of PCK which appears to be unclear in current literature.