L-arginine is an alkaline amino acid that has been used as a neutralizer, moisturizer, and antioxidant in skin care products. In addition, L-arginine is also widely used in feed, medicine, and food industries. The wide range of applications for L-arginine has garnered significant attention for its robust production. L-arginine can be produced through protein hydrolysis and microbial fermentation. However, protein hydrolysis has drawbacks, including complicated operation, high purification cost, low recovery efficiency, and environmental pollution. In contrast, the microbial fermentation can use renewable and cheap feedstock. Besides, the process is performed under mild conditions, and thus is more environmentally friendly. At present, engineered microorganisms such as Corynebacterium glutamicum and Escherichia coli are major producers of L-arginine, and design and construction of microbial strains is the robust production of L-arginine through microbial fermentation. Random mutagenesis and screening strategies are used to develop L-arginine producing microbial strains, which are random with uncertainties, resulting in a low-efficiency for the breeding. With the development of synthetic biotechnology, development of L-arginine producing strains is empowered by the rational design of artificial synthetic pathways and regulatory machineries, taking advantages of advanced genome editing technologies. This paper reviews the progress in the studies of the synthetic pathways and regulatory mechanisms of L-arginine production that have been discovered in different microorganisms. Synthetic biology-guided metabolic engineering strategies for improving L-arginine production in C. glutamicum and E. coli are summarized. Besides, the application of the biosensor-based high-throughput screening strategy for selecting L-arginine producing strains is introduced. Finally, potential strategies to enhancing L-arginine production and the possibility of using new carbon resources such as non-food biomass and one-carbon feedstock for L-arginine production are discussed. It is envisioned that synthetic biology-guided strain engineering will further enhance the production of L-arginine, particularly using non-food feedstock in the near future.