Synthetic biology has led to the creation of new products, markets, companies, and industries. At the same time, this technology poses potential risks to biosafety and biosecurity, as recently demonstrated by the synthesis of horsepox virus, a cousin of variola, the virus that causes smallpox. On January 29-30, 2018, the Department of Homeland Security (DHS) Science and Technology Directorate sponsored a workshop to discuss the evolving role of databases that contain genetic sequences of pathogens and toxins that pose safety or security concerns, termed “sequences of interest.” The workshop brought together stakeholders from government, industry, and academia to discuss the need for such databases, review current databases and those under development, explore potential applications and users of these types of databases, and consider the potential risks that they pose due to malicious or inadvertent misuse. The workshop provided a valuable opportunity to explore the scientific and technical aspects of constructing such databases, maintenance and sustainability challenges, and the trade-offs involving functionality, accessibility, affordability, confidentiality, and security. While the workshop did not produce a consensus on the best path forward, it played an important role in educating the participants on the most critical issues and facilitating a dialogue among a diverse range of stakeholders on this important topic. The workshop also came at a propitious time as stakeholders grapple with the changing landscape of the biotech industry and advances in DNA synthesis technology. For instance, the International Gene Synthesis Consortium, a group of the leading DNA synthesis companies that have adopted customer and sequence screening protocols to prevent the misuse of their products, has expanded in size and geographic scope and recently updated its biosecurity protocols. The U.S. government is also reviewing the customer and sequence screening guidance it issued in 2010 and is considering whether, and how, to update it. Should a decision be made to proceed, stakeholder engagement would be a significant part of the review, just as it was a significant part of the guidance’s original development.