Abstract Great strides are continually made in showing how the initiation of individual replication forks is regulated, and the next frontier is to understand how DNA replication is coordinated with transcription and chromatin structure. In particular, little is known about the mechanisms and function of DNA replication control during vertebrate development, when transcription and chromatin structure are highly dynamic. This gap in knowledge is an important problem because, until it is filled, the roles for DNA replication in developmental disorders and cancers associated with epigenetic or DNA replication deregulation will be largely incomprehensible. With the unique capability of using both traditional cell culture as well as zebrafish embryos as model systems, we are uniquely poised to define the key factors in the regulation of replication fork initiation. In this application, we are requesting funds through an equipment supplement to purchase a replacement biosafety cabinet that can be used with the Aims of NIH GM121703. The existing biosafety cabinet is over 35 years old and has become obsolete. All experiments in Aim 3 require the use of a BSL2 biosafety cabinet. This Aim tests whether early replication of acetylated chromatin depends on the physical interaction of an essential replication factor, TICRR, and the epigenetic reader protein BRD4. Results from this aim show that disruption of the TICRR-BRD interaction leads to aberrant timing of replication initiation during S-phase. To address how specific replication initiation proteins ensure that large genomic segments are replicated at the correct time, engineered endogenously tagged cell lines of additional replication factors will be used. Successful achievement of the research in GM121703 is significant because it seeks to answer fundamental questions about how and why DNA replication initiation changes throughout development.