The development of fields such as bioinformatics and genomics has created new challenges and opportunities for undergraduate biology curricula. Students preparing for careers in science, technology, and medicine need more intensive study of bioinformatics and more sophisticated training in the mathematics on which this field is based. In this study, we deliberately integrated bioinformatics instruction at multiple course levels into an existing biology curriculum. Students in an introductory biology course, intermediate lab courses, and advanced project-oriented courses all participated in new course components designed to sequentially introduce bioinformatics skills and knowledge, as well as computational approaches that are common to many bioinformatics applications. In each course, bioinformatics learning was embedded in an existing disciplinary instructional sequence, as opposed to having a single course where all bioinformatics learning occurs. We designed direct and indirect assessment tools to follow student progress through the course sequence. Our data show significant gains in both student confidence and ability in bioinformatics during individual courses and as course level increases. Despite evidence of substantial student learning in both bioinformatics and mathematics, students were skeptical about the link between learning bioinformatics and learning mathematics. While our approach resulted in substantial learning gains, student "buy-in" and engagement might be better in longer project-based activities that demand application of skills to research problems. Nevertheless, in situations where a concentrated focus on project-oriented bioinformatics is not possible or desirable, our approach of integrating multiple smaller components into an existing curriculum provides an alternative. (Contains 4 tables.)