RNA, a crucial molecule in protein synthesis and gene expression regulation, plays an essential role in organisms. RNA modifications, acting as epigenetic marks, subtly adjust the structure, stability, and function of RNA, thereby regulating gene expression and exerting profound effects on cellular functions and organismal health. These natural modifications, together with RNA editing that alters nucleotide sequence of mRNA, constitute the epitranscriptome, which is vital for cellular metabolism. With the rapid advancement of biotechnology, RNA-based therapies and technologies have emerged as a frontier in biotech research. Various RNA drugs, including small interfering RNA (siRNA), antisense oligonucleotides (ASO), mRNA vaccines, and the small guide RNA (sgRNA) required for CRISPR gene editing, have been continuously developed. It has been widely demonstrated that RNA modifications can alter the physicochemical properties of RNA, enhance resistance to nucleases, reduce immunogenicity, and optimize in vivo functionality, leading to their extensive application in RNA-related biotechnologies. Furthermore, the variety of RNA modifications has expanded beyond natural modifications with the invention of an increasing number of artificial modifications. This review delves into the common types of RNA modifications, including base, ribose, and phosphate modifications, discussing their impact on RNA structure and how these modifications influence the biological characteristics of RNA. The current applications of these modifications in the biotechnology field are summarized, highlighting their significance in RNA-based therapies.