Over the past decades, the development of microbial cell factories for industrial applications was limited by the over-reliance on non-optimal model organisms. These non-optimal hosts are applied in bioproduction processes or for bioengineering. However, typically cell resources are used inefficiently, and there is a toxic build-up of intermediates, causing a metabolic burden and leading to lower yields and suboptimal processes. Indeed, a shift towards microorganisms more suited for the application requirements is a possible solution to this problem. Unfortunately, the lack of orthogonal and standardized genetic parts for alternative hosts is a significant bottleneck for the predictable engineering of these hosts. To address this development gap, this project aims to develop a novel orthogonal synthetic biology toolbox dedicated to thermophilic bacteria, using bacteriophages as synthetic biology part suppliers. Furthermore, the orthogonality of the parts further minimizes any possible metabolic burden on the thermophilic host. The development of these new parts allows a new paradigm in industrial biotechnology and opens infinite possibilities. To accomplish this, bacteriophages associated with thermophiles are in silico mined to harvest new biological DNA parts. Harvested parts are characterized by functionality and orthogonality, followed by tuning and optimization. Eventually, novel synthetic circuits are designed, developed and evaluated for thermophilic hosts.