Synthetic plastics have been widely used in various fields of the national economy and are the pillar industry. However, irregular production, plastic product use, and plastic waste piling have caused long-term accumulation in the environment, contributing considerably to the global solid waste stream and environmental plastic pollution, which has become a global problem to be solved. Biodegradation has recently emerged as a viable disposal method for a circular plastic economy and has become a thriving research area. In recent years, important breakthroughs have been made in the screening, isolation, and identification of plastic-degrading microorganisms/enzyme resources and their further engineering, which provide new ideas and solutions for treating microplastics in the environment and the closed-loop bio-recycling of waste plastics. On the other hand, the use of microorganisms (pure cultures or consortia) to further transform different plastic degradants into biodegradable plastics and other compounds with high added value is of great significance, promoting the development of a plastic recycling economy and reducing the carbon emission of plastics in their life cycle. We edited a Special Issue on the topic of "Biotechnology of Plastic Waste Degradation and Valorization", focusing on the researches progress in three aspects: Mining microbial and enzyme resources for plastic biodegradation, Design and engineering of plastic depolymerase, and biological high-value transformation of plastic degradants. In total, 16 papers have been collected in this issue including reviews, comments, and research articles, which provide reference and guidance for further development of plastic waste degradation and valorization biotechnology.