Synthetic biology has garnered much attention in recent years due to its wide number of applications in almost every field of biological sciences, which engineer the living systems for one's own benefit. Although human history has witnessed the use of living cells for the production of alcohol, enzymes, antibiotics, food supplements, etc. However, the manipulation of DNA, protein, or metabolic pathway of the living systems is now the latest research interest. There are now biological counterparts of almost every chemical or synthetic substance available such as biopolymers, biofertilizers, biorubbers, bioplastics, and many more that may be credited to synthetic biology systems. The list of areas profiting from synthetic biology systems ranges from food, drug, agriculture industries to bioenergy, environment, etc. This chapter explores the most recent research works, tools, technologies, and applications of synthetic biology systems in the area of enzymes, secondary metabolites production, pharmaceuticals, biofuels, and cell-free protein synthesis.